How are antibodies extracted from donor blood?

How are antibodies extracted from donor blood?

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When blood is donated, the antibodies within it are extracted, but how exactly do they do it? How do they take out the antibodies within the blood, what process do they go through?

I don't believe they do remove the antibodies but rather they match and screen the donor antigen and antibody profiles to those of the recipient. The process is very briefly outlined on the Red Cross website;

  1. Most blood is spun in centrifuges to separate the transfusable components - red cells, platelets, and plasma

  2. The primary components like plasma, can be further manufactured into components such as cryoprecipitate

  3. Red cells are then leuko-reduced

  4. Single donor platelets are leukoreduced and bacterially tested.

The article in wikipedia explains the screening for blood transfusions;

Patients should ideally receive their own blood or type-specific blood products to minimize the chance of a transfusion reaction. Risks can be further reduced by cross-matching blood, but this may be skipped when blood is required for an emergency. Cross-matching involves mixing a sample of the recipient's serum with a sample of the donor's red blood cells and checking if the mixture agglutinates, or forms clumps. If agglutination is not obvious by direct vision, blood bank technicians usually check for agglutination with a microscope. If agglutination occurs, that particular donor's blood cannot be transfused to that particular recipient.


An antibody is a specialized defense protein synthesized by the vertebrate immune system. These small structures are actually made of 4 different protein units. The ends of the molecule are variable, and can be adapted to bind to any molecule. The shape is determined by the antigens in the system which are causing damage. Special immune cells detect these antigens and create a reciprocal antibody. This generalized structure is repeated many times, to flood the system with antibodies. These proteins bind to and surround the antigens, preventing further spread or infection.

It is in this way that an organism can identify “self” from “non-self”. For instance, the surface of bacterial cells has certain proteins and carbohydrates, which can be identified by the immune system. B lymphocytes, a special immune cell, create and release antibodies which attack the invading bacteria. An antibody attached to a bacteria not only prevents it from completing normal processes, but helps direct white blood cells to eat the bacteria. These macrophages, as they are known, identify food based on the tail end of the antibody.

In the blood, antibodies account for around 20% of the total protein. This is a very significant amount. Although a single antibody can be very small, an organisms must have many antibodies to fight the many types of antigens present in the system. Further, many of each type is needed. It often takes many antibody molecules to target and identify a large bacteria. Viruses, while they are smaller, are much more abundant and need equal amounts of antibody to quell.

While other organisms often have immune systems based on similar concepts, the term antibody and the structure described below are unique to mammals. An antibody may also be referred to as immunoglobulin, a term which describes a protein used in an immune function. The most common antibody is Immunoglobulin G (IgG) in mammals. Antibodies, if they exist, are not well understood in invertebrates and plants. While it is known that these organisms also have immune systems, it is not entirely clear how they function.

Chronic graft-versus-host disease (cGVHD) after allogeneic hematopoietic stem cell transplant reflects a complex immune response resulting in chronic damage to multiple tissues. Previous studies indicated that donor B cells and the antibodies they produce play an important role in the development of cGVHD. To understand the pathogenic role of antibodies in cGVHD, we focused our studies on posttransplant production of immunoglobulin G antibodies targeting cell surface antigens expressed in multiple cGVHD affected tissues, due to their potential functional impact on living cells in vivo. Using plate-bound cell membrane proteins as targets, we detected a significantly higher level of antibodies reactive with these membrane antigens in patients who developed cGVHD, compared with those who did not and healthy donors. Plasma-reactive antibody levels increased significantly prior to the clinical diagnosis of cGVHD and were reduced following cGVHD therapies including prednisone, interleukin-2, or extracorporeal photophoresis. Using cell-based immunoprecipitation with plasma from cGVHD patients and mass spectrometry, we identified 43 membrane proteins targeted by these antibodies. The presence of antibodies in cGVHD patients’ plasma that specifically target 6 of these proteins was validated. Antibodies reactive with these 6 antigens were more frequently detected in patients with cGVHD compared with patients without cGVHD and healthy donors. These results indicate that antibodies that target membrane antigens of living cells frequently develop in cGVHD patients and further support a role for B cells and antibodies in the development of cGVHD.

Chronic graft-versus-host disease (cGVHD) is a major complication of allogeneic hematopoietic stem cell transplant (HSCT) and remains a leading cause of long-term morbidity and mortality. 1-4 The clinical manifestations of cGVHD can affect multiple organ systems and often resemble autoimmune diseases. 3 Recent studies in murine models and patients have provided new insights into the complex immunopathologic mechanisms that contribute to immune dysregulation and the development of cGVHD months to years after allogeneic HSCT. 4,5 These studies highlight the complexity of immune responses in cGVHD and interactions between adaptive and innate immune systems resulting in chronic inflammation and tissue damage. 6

In this context, many studies have demonstrated an important role for B-cell immune responses in cGVHD. Donor B cells can function as antigen-presenting cells to facilitate the development of cGVHD. 7-10 In these studies, antigen presentation by donor B cells promotes the survival and proliferation of pathogenic CD4 + T cells and is essential for the development of cGVHD caused by donor T cells. In addition to their antigen-presentation function, recent studies revealed that the antibodies produced by B cells also play an important role in the pathogenesis of cGVHD. Antibodies against HY minor histocompatibility antigens frequently develop in male recipients with female stem cell donors and the development of these antibodies is significantly increased in patients with cGVHD. 11-13 Notably, anti-HY antibodies can be detected as early as 3 months after transplant and precede the clinical onset of cGVHD. 14 In murine models, antibodies produced by donor B cells are required for the development of cGVHD. Immunoglobulin G (IgG)-containing donor serum leads to deposition of IgG in thymus and skin, resulting in tissue damage and perpetuation of cGVHD. Interventions that prevent B-cell recovery or inhibit B-cell function prevent or delay the onset of cGVHD. 15-17

In addition to antibodies against minor histocompatibility antigens, studies have identified antibodies specific for other antigens in patients with cGVHD. However, most antibodies previously identified in this setting are specific for intracellular antigens. 11,18,19 Considering the important role that antibodies play in the development of cGVHD, we hypothesized that patients with cGVHD also develop antibodies that target surface antigens on live cells. Antibodies directed against surface membrane antigens would have a direct functional impact on target cells in affected tissues and could help explain the mechanism whereby antibodies contribute to the pathogenesis of cGVHD. To address this issue, we developed specific assays to monitor the development of antibodies that target membrane antigens after allogeneic HSCT. We further investigated the specific targets of these anti-membrane antibodies and validated the specificity of antibodies against 6 membrane antigens in patients with cGVHD. Our goals were to characterize the generation of anti-membrane antibodies in cGVHD and correlate the levels of anti-membrane antibodies with the onset of cGVHD and response to immune-suppressive therapy.

Why don't a blood donor's antibodies cause problems for the reciever?

Blood typing is always done to make sure the reciever's body doesn't reject the blood because it has antibodies against it.

But what about the donor? Why is it okay for an A-type, who has anti B antibodies to donate their blood to an AB-type? Or an O who has antibodies for everyone, how are they a universal donor?

2 2

When whole blood is donated, it’s separated into its different components (red cells, plasma, and platelets). The anti-B antibodies in a type A person are only in the plasma. So, giving A red cells to an A person or an AB person is safe because it’s only the red cells. The same goes for O blood. It’s universal because it’s just the O cells. A plasma can only go to A and O people because they’re the only ones who can handle anti-B antibodies. O plasma can only go to O patients because it does have anti-A and anti-B. Does that make sense?


Donor T Cells Are the Most Effective Inducers of the DST Antibody Response

When fresh blood of ACI rats was transfused to Lewis rats, an equivalent number of purified T cells and WBC fractions present in the blood induced a comparable level of DST antibodies ( Figure 1A ). Flow cytometric analysis (FCM) of recipient spleen cells revealed similar proliferative responses of CD4 + T cells, Foxp3 + regulatory T cells, and CD45R + B cells as in those who received whole blood ( Figure 1B ). In contrast, transfusion of other components, such as purified RBCs, platelets, or blood plasma, induced a significantly delayed and less intense response ( Figures 1A,B ). Furthermore, when T cells were depleted from the WBC fractions, the serum DST antibody response was suppressed considerably at 7 days ( Figure 1C ).

Screening of effective blood components for the donor-specific transfusion (DST) response. (A) Production of DST antibodies in recipient blood after donor blood component injection. In the white blood cell (WBC) and T cell injected groups, DST antibodies were detected after 5 days (mean ± SD, n = 3 rats each, *P < 0.05). (B) The time kinetic changes in each proliferating cell type as a proportion of total splenocytes. Two-color FCM analysis of recipient splenic lymphocytes after donor blood component injection for proliferating cells (EdU + ) and CD4 + T cells, CD8β + T cells, CD45R + B cells, or regulatory T cells (Foxp3 + ). The proliferation of CD4 + T cells and regulatory T cells peaked 3 days after treatment, whereas B cells and CD8β + T cells peaked at 5 days. These results were similar to the DST group that received whole blood (mean ± SD, n = 3 rats each, *P < 0.05). (C) With T cell depletion of WBCs, the DST antibody production was suppressed at 7 days. (D) Dose-dependent response following injection of isolated T cells or B cells. T cells induced much higher DST antibody production than B cells (mean ± SD, n = 3 rats each, *P < 0.05). MFI, mean fluorescent intensity.

For dose response, we employed thoracic duct lymphocytes (TDLs) as a convenient source of donor lymphocytes, which were functionally almost the same as those in the blood (25, 26). At least 3 × 10 5 T cells from donor TDLs, representing those in

100 μl of blood, induced significant DST antibody production ( Figure 1D ). In contrast, 10 6 B cells induced a significant but very low level comparable to that induced by 1 × 10 5 T cells ( Figure 1D ). When equivalent numbers of purified CD4 + T cells and CD8 + T cells ware transferred, both induced comparable DST antibody production (Figure S1), indicating both subsets are equally efficient.

These results indicate that T cells are the most proficient immunogens for the DST response among blood components. A pure T cell fraction, both CD4 + and CD8 + , without DCs or monocytes (equivalent to

100 μl blood) is enough for significant induction of a DST antibody response in the ACI to Lewis rat combination, and B cells require

Fate of Donor Cells in the Spleen and the LNs

Immunohistological examination of recipient spleens found that, 1 day after DST, donor MHCI + cells were found in the splenic T cell area, the periarterial lymphocyte sheath (PALS) ( Figure 2A ), which consisted mainly T cells (71.1 ± 3.4%) and B cells (27.9 ± 7.8%) but few other cell types ( Figure 2B ). This migration was very fast and readily observed by 15 min after DST, as reported previously (27).

Fate of donor T cells after DST. Three-color immunostaining of recipient spleens (A𠄽) or peripheral LNs (E–G) for donor MHCI (blue), BrdU (red), and type IV collagen (brown) or recipient MHCII (brown). (A,E) Donor MHCI + cells (arrows) were observed in the PALS (A) and T cell area of the LNs (paracortex, PC) (E) at day 1. (B) Phenotype of donor MHCI + cells and cell ratio in the PALS. Approximately 100 donor MHCI + cells/rat were examined (n = 3). Donor MHCI + cells that migrated to the PALS were TCRαβ + T cells (

30%). (C,F) A donor cell binding to MHCII + putative recipient DCs (brown) became BrdU + (arrowhead) in the PALS (C) and the PC (F) at day 1, representing a graft vs. host reaction. (D,G) Donor MHCI + fragments (blue) superimposed on the cytoplasm of recipient putative DCs (arrows) in the PALS (D) and the PC (G) at day 2, suggesting phagocytosis. F, lymph follicle P, splenic PALS Z, marginal zone HEV, high endothelial venule. Scale bar = 100 μm (A,E), 20 μm (C,F), or 10 μm (D,G).

Many of migrated T cells bound to recipient class II MHC (MHCII) + cells with dendritic cytology in the PALS, and some became large in size and BrdU + ( Figure 2C ). As these MHCII + cells are mostly CD103 + CD11b + resident interdigitating DCs (14), the result indicates a graft vs. host (GvH) reaction via the direct pathway in which donor T cells become activated within the cluster with recipient DCs (14, 27).

Donor MHCI + cells decreased in number at 2 days and almost disappeared by 3 days. From day 1, donor MHCI + fragments appeared in the PALS, and at day 2 they often superimposed on the cytoplasm of some recipient MHCII + cells with dendritic cytology ( Figure 2D ). This suggests phagocytosis of donor MHCI + fragments by recipient DCs in the PALS.

As for the LNs, donor MHCI + cells also readily migrated to the T cell area of LNs, paracortex, and showed almost the same kinetics as that in the spleen ( Figures 2E–G ).

Donor T Cells Fragments Are Phagocytosed by XCR1 + Resident DCs in the PALS

To confirm phagocytosis of donor MHCI fragments and the phenotype of phagocytic DCs, we examined subsets of conventional DCs that were CD103 + MHCII + in the normal rat spleen (28, 29). FCM detected two subsets, CD4 − XCR1 + signal regulatory protein 1 α (SIRP1α) − CD200 + cells and CD4 + XCR1 − SIRP1α + CD200 − cells (Figure S2A). SIRP1α is CD172a. Because mouse CD8 + DCs were recently defined as XCR1 + SIRP1α − (30, 31), it is reasonable to consider the CD4 − XCR1 + SIRP1α − CD200 + subset as the rat counterpart of mouse CD8 + DCs. These subsets were detected by immunohistology in the PALS and a CD103 + XCR1 + subset constituted 40.1 ± 6.9% in a normal steady state (Figures S2C,E).

In mice, XCR1 are exclusively expressed by CD8 + DC subset (31). In rats, however, the mAb to XCR1 reacted with not only DCs but also CD103 − non-DCs (Figure S2C). Double immunostaining of rat spleens for XCR1 and a macrophage marker CD169 (Figure S2D) or CD163 (not shown) revealed a presence of XCR1 + CD169 + cells and XCR1 + CD163 + cells in the marginal zone and red pulp, respectively. CD169 + cells in the outer margin of the PALS and in the marginal zone are marginal metallophilic macrophages and marginal zone macrophages, respectively and CD163 + cells are red pulp macrophages (27, 32). The mAb we used exclusively reacted with mice DCs (not shown), while isotype control of the XCR1 mAb (IgG2b) did not detect CD103 + DCs or CD169 + and CD163 + macrophages (Figure S2D), confirming the specificity of the mAb. Accordingly, the results indicate that at least some macrophages in the rat spleen also express XCR1.

Next, we examined isolated spleen cells and cryosections 36 h after transfer of CFSE-labeled donor TDLs ( Figure 3A ). FCM revealed that CD103 + fractions contained CFSE + signals, and that these cells were CD103 + MHCII high XCR1 + SIRP1α − DCs ( Figure 3B ). When the CFSE + CD103 + MHCII high DC fraction was sorted and cytosmeared, these DCs were XCR1 + SIRP1α − and contained cytoplasmic donor CFSE + fragments ( Figure 3C ), demonstrating phagocytosis of donor cell fragments.

Phagocytosis of donor MHCI fragments and phenotype of phagocytic DCs in the spleen. (A) Experimental protocol for identifying DCs that phagocytosed donor cells. (B) Three-color FCM analysis of magnetically isolated CD103 + DC fraction of the recipient spleen 36 h after CFSE-labeled thoracic duct lymphocyte (TDL) injection for CFSE, XCR1, or SIRP1α, and recipient MHCII. XCR1 + SIRP1α − CD103 + MHCII high DCs expressing CFSE + signals (red dots) were detected within the gated fraction (1.05%) of the SSC vs. CFSE plot. A representative data of 4 individual experiments. (C) Two-color immunofluorescent staining of cytosmears of sorted CD103 + recipient MHCII high CFSE + cells for DAPI (DNA stain, blue), and XCRI or SIRP1α (red). CFSE + donor cell fragments (green) were observed in the cytoplasm of XCR1 + SIRP1α − DCs but not XCR1 − SIRP1α + DCs, indicating phagocytosis. Scale bar = 10 μm. (D) Three-color immunofluorescent staining of the PALS cryosections for donor MHCI (green), CD103 (blue), and XCR1 (red). The arrowheads indicate XCR1 + CD103 + DCs with phagocytosed donor MHCI + fragments. Scale bar = 10 μm. (E) The subset and ratio of CFSE + phagocytic cells in different splenic areas. In the PALS, XCR1 + CD103 + DCs accounted for more than 80% in CFSE + phagocytic cells (mean ± SD, n = 4 rats each). In contrast, more than 90% phagocytic cells in the marginal zone (MZ) and red pulp (RP) were XCR1 + CD103 − non-DCs.

Of note, the XCR1 + DCs could be separated into two populations according to their phagocytic activities and phenotype, where phagocytic cells were mostly XCR1 low

int MHCII high CD103 + and non-phagocytic cells were mostly XCR1 high MHCII int CD103 + , respectively ( Figure 3B ). We do not know whether this difference is due to phagocytic events or not, because untreated rat spleen also contained these two populations (Figure S2B).

Immunohistology revealed donor MHCI + fragments superimposed on CD103 + XCR1 + DCs in the PALS ( Figure 3D ), representing phagocytosis. When the phenotypes of phagocytic cells in the different components of the spleen were examined, more than 80% of the cells in the PALS were CD103 + XCR1 + DCs ( Figure 3E ). Although CD103 + XCR1 + or CD103 + XCR1 − DCs were also present in the marginal zone and the red pulp, phagocytosis of the donor MHCI + fragments was negligible ( Figure 3E ).

These data demonstrate that XCR1 + SIRP1α − MHCII high CD103 + resident DCs selectively phagocytose donor cell fragments in the PALS and that XCR1 + macrophages outside of the PALS also phagocytose them as scavengers.

NK Cells Are Responsible for Donor Cell Phagocytosis in the PALS

The number of asialo GM 1 + NK cells/mm 2 of PALS significantly increased 4 h after DST ( Figures 4A𠄼 ). After NK cell depletion with anti-asialo GM1 antibody, CD103 + XCR1 + DCs ingesting CFSE + donor T cell fragments significantly decreased in number ( Figures 4D𠄿 ). The production of serum DST antibodies 7 days after DST was suppressed almost completely ( Figure 4G ). Although we performed quantitative PCR (qPCR) analysis of the whole spleen shortly after the donor T cell transfer, we could not detect increased mRNA levels of NK-recruiting chemokines (Figure S3A), probably because the increase in NK cell number was small,

Kinetics of NK cells and donor cell phagocytosis in the PALS. Double immunostaining for asialo GM1 (blue) and rat IgM (brown) in the splenic PALS 0 (A) and 4 h (B) after donor-specific transfusion (DST). Arrows indicate asialo GM 1 + NK cells. (P, splenic PALS. Scale bar = 100 μm. (C) Number of asialo GM 1 + NK cells/mm 2 of PALS 0, 4, and 12 h after DST. NK cells significantly increased in the PALS 4 h after DST. Data was analyzed by Mann-Whitney U-test. (D) Experimental protocol for examining the effect of NK cell depletion by anti-asialo GM1 antibody (Ab) on the DST response. PTX, pertussis toxin. (E,F) With NK cell depletion, DCs ingesting CFSE + donor T cell fragments in the spleen significantly decreased in number to the same level of the syngeneic combination group. PTX pretreatment of donor T cells also resulted in the significant decrease. mean ± SD, n = 3 rats each, *P < 0.05. allo, allogeneic ACI to Lewis syn, syngeneic Lewis to Lewis. (G) In the NK cell-depleted group, the DST antibody response disappeared completely (mean ± SD, n = 3 rats each, *P < 0.05). MFI, mean fluorescent intensity.

Concerning NK activity against allogeneic cells (33), other two rat strains, PvG/c and BN rats, which readily showed the DST antibody response by 7 d after DST treatment, were compared as recipients (Figures S4A,B). ACI rats were used as donors. PvG/c rats clearly exhibited accelerated phagocytosis by XCR1 + DCs, as it occurred 1 day earlier than with Lewis rats (Figures S4C,G). When BN rats were used as recipients, donor T cells (Figure S4D) extensively proliferated (Figure S4E), indicating a predominance of GvH reaction, probably due to low NK activity. Because this causes dilution of the CFSE + signals of donor T cells, detection of phagocytosis becomes difficult. Accordingly, we transferred mitomycin C (MMC)-treated donor T cells to inhibit their proliferation. With this, donor T cells disappeared (Figure S4F) and we readily observed phagocytosis at 1.5 days (Figure S4H). These results indicate that NK activities correlated well with the phagocytosis by XCR1 + DCs in at least three rat strain combinations (Figure S4I). As strain differences in NK reactivity is a rather complex issue, other mechanisms of donor cell killing may be involved in other rat strain combinations.

Alternatively, CD161a + (NK1.1 + ) killer DCs have been reported in the rat spleen (28) and effector/memory fraction of CD8 + T-cells is known to be asialo GM 1 + in mice (34). These cells may be related to donor cell phagocytosis. Although two subsets of splenic DCs were both CD11b + CD161a + , they were asialo GM 1 - (Figure S3C) and not depleted by anti-asialo GM1 antibody (data not shown). Also, we found that in rats, CD8 + T-cells and TCRαβ + CD161a + putative NKT cells were mostly asialo GM 1 - (Figure S3D). These findings suggest that rat NK cells are mostly asialo GM 1 + CD161a + . Because asialo GM 1 + cells are either CD8α + or CD8α − (Figure S3D), NK cells can be further divided into two subsets. In fact, anti-CD8α monoclonal antibody (mAb) treatment resulted in only partial depletion of NK cells (35) and partial suppression of phagocytosis (unpublished data, Ueta). Therefore, the results indicate that involvement of killer DCs or CD8 + T-cells would be minor for the donor cell phagocytosis.

Migration of Donor T Cells to the PALS Is Crucial for the DST Response

To inhibit donor T cell migration to the PALS, we blocked the chemokine-chemokine receptor axis with pertussis toxin (PTX). Compared to the readable migration of untreated cells into the PALS, PTX-treated T cells did not enter the PALS for up to 3 days, staying in the marginal zone and red pulp for 1 day and soon disappearing ( Figure 5A ). In this group, DCs ingesting CFSE + donor cell fragments in the spleen significantly decreased in number ( Figures 4E,F ), and the DST responses of CD4 + T cells and CD45R + B cells ( Figure 5B ) and serum antibodies ( Figure 5C ) were abrogated. These results indicate that the migration of donor T cells into the PALS is indispensable for induction of the DST response.

Requirement of T cell migration to the PALS for induction of the DST response. (A) Two-color immunostaining for donor MHCI (blue) and type IV collagen (brown) in the spleen at 1 and 3 days after 2 × 10 6 donor T cell injection and in the peripheral LNs at 18 h after 2 × 10 7 donor T cell injection. The control group exhibited migration to the PALS by 1 day (Cont T 1d, arrowheads) and LNs by 18 h (Cont T 18 h, arrowheads). In contrast, pertussis toxin (PTX) pretreated donor T cells did not enter the PALS up to 3 days (PTX T 3d), but stayed in the marginal zone and red pulp for 1 day (PTX T 1d, arrows). They did not enter the LNs (PTX T 18 h) either. F, lymph follicle HEV, high endothelial venule P, PALS PC, paracortex. Scale bar = 100 μm or 20 μm (inset). (B,C) With PTX pretreatment of donor T cells, the DST responses of CD4 + T cells and CD45R + B cells (B) and serum antibodies (C) were abrogated (mean ± SD, n = 3 rats each, *P < 0.05). MFI, mean fluorescent intensity.

Donor T Cells Also Induced the DST Response in the LNs

To avoid the influence of the spleen on DST responses in the LNs, rats were splenectomized just before donor T cell transfer. Donor T cells migrated to all of the examined LNs, and their number was approximately double that of eusplenic recipients ( Figure 6A ). The peripheral LNs ( Figures 6B,C ) and gut LNs (data not shown) demonstrated proliferative responses of recipient T and B cells with similar kinetics as those observed in the spleen ( Figure 1B ). NK cells significantly increased by 12 h ( Figures 6D,E ), and phagocytosis of donor cell fragments by XCR1 + DCs was detected in LN cell suspensions ( Figures 6F,G ). PTX pretreatment also suppressed donor T cell migration to the LNs ( Figure 5A ) and inhibited the DST antibody response (data not shown). Anti-asialo GM1 antibody pretreatment significantly suppressed the NK cell number and phagocytosis by XCR1 + DCs ( Figure 6F ). However, we did not detect increased mRNA levels of NK-recruiting chemokines in LN extracts (Figure S3B).

Induction of the donor-specific transfusion (DST) response in the LNs of splenectomized (A𠄼,F,G) or eusplenic (D,E) recipients. The recipient peripheral LNs were analyzed in the same manner as the spleen ( Figure 1 , ​ ,3, 3 , ​ ,4). 4 ). (A) Two-color FCM analysis of the number of migrated donor T cells in recipient LNs 24 h after injection. Roughly twice as many donor T cells migrated to the LNs than in the eusplenic control. SPL, spleen CLN, cervical LN BLN, brachial LN ALN, axillary LN HLN, hepatic LN MLN, mesenteric LN. (B,C) The time kinetic changes in each proliferating cell type/total LN cells by FCM analysis. The CD4 + T cells and regulatory T cells peaked at 3𠄵 days and 3 days, respectively, whereas CD45R + B cells and CD8β + T cells peaked at 5 days (mean ± SD, n = 3 rats each, *P < 0.05). (D,E) Kinetics of NK cells in the peripheral LNs after DST. (D) Double immunostaining of the LNs for asialo GM1 (blue) and rat IgM (brown) at 0 (left panel) and 12 h (right panel). Inset indicates increased asialo GM 1 + NK cells (blue) in the paracortex (PC). Scale bar = 200 μm or 50 μm (inset). F, lymph follicle. (E) The proportion of NK cells significantly increased in the LNs at 12 h (mean ± SD, n = 3 rats each, *P < 0.05). (F) Kinetics of donor cell phagocytosis in the peripheral LNs after donor T cell injection. Phagocytosis of donor cell fragments by DCs was detected 36 h after transfer of 2 × 10 7 CFSE labeled T cells. With anti-asialo GM1 antibody (Ab) treatment, the numbers of recipient CFSE + phagocytic DCs and NK cells were significantly reduced compared to non-treated control (mean ± SD, n = 3 rats each, *P < 0.05). (G) Phenotype of CFSE + phagocytic DCs in the peripheral LNs. Note that they were XCR1 + SIRP1α − , similar as those of spleen.

These results indicate that allogeneic T cells induced a DST antibody response in the peripheral LNs in a similar manner as in the spleen, dependent on the NK response and phagocytosis by XCR1 + DCs in ACI to Lewis rat combination.

GvH Reaction Is Not Required for the DST Response

Because donor T cells exhibited a GvH reaction at 1𠄲 days ( Figures 2C,F ), we examined whether donor T cell activation is required for the DST response. T cells from (Lewis × DA)F1 hybrid rats (RT1.A al B al ) were transferred to parental Lewis rats (RT1.A l B l ), a semiallogeneic combination in which the GvH reaction does not occur. The donor T cells induced significant serum DST (anti-RT1.A a ) antibody production on day 7 (Figure S5). Furthermore, fully allogeneic donor T cells pretreated with MMC, migrated to the SLOs and induced DST antibody production as will be described below. Thus, the GvH reaction leading to donor T cell activation and proliferation is dispensable for the DST response.

Both XCR1 + and XCR1 − DCs Form a Cluster With Proliferating Recipient T Cells

When activation state of DC subsets was examined 36 h after transfer of CFSE-labeled donor TDLs, CFSE + XCR1 + DCs showed a significant upregulation of CD40 ( Figures 7A,B ), although CFSE − XCR1 + and CFSE − XCR1 − DCs showed no change (Figure S6). CD103 + DCs readily formed clusters with proliferating cells in the PALS 2 days after DST ( Figures 7C,D ), which was composed mostly of CD4 + T cells as reported previously (24). Although we observed selective phagocytosis of donor cells by XCR1 + DCs ( Figure 3 ), not only XCR1 + DCs (

16%) formed clusters with recipient EdU + proliferating cells in the PALS ( Figures 7E,F ). Cluster-forming XCR1 + CD103 − non-DCs were also found ( Figure 7F ) and may be a macrophage lineage.

Activation state of phagocytic dendritic cells (DCs) and phenotype of DCs that cluster with proliferating cells in the PALS. (A,B) Upregulation of CD40 in XCR1 + DCs following phagocytosis. Phagocytic DCs were compared with non-phagocytic DCs in MHCII high gates with/without donor cell transfer (mean ± SD, n = 4 rats each, *P < 0.05, NS, not significant). (C,D) Three-color immunostaining of the spleen 2 days after DST for CD103 (blue), BrdU (red), and type IV collagen (brown). (C) Many CD103 + DCs (blue) and BrdU + proliferating cells (red) are present in the PALS (P). Scale bar = 100 μm. F, lymph follicle Z, marginal zone. (D) Inset of A showing cluster formation (arrowheads) of recipient DCs (blue) with BrdU + cells (red). Scale bar = 20 μm. (E) Three-color immunofluorescence staining of the PALS for XCR1 (green), CD103 (red), and EdU (white) 2 days after DST, showing clusters (arrowheads) of XCR1 + CD103 + (left panels) or XCR1 − CD103 + (right panels) DCs with EdU + proliferating cells (white). Scale bar = 10 μm. (F) Proportion of cluster-forming cells in the PALS, showing

35% were XCR1 + CD103 + DCs, XCR1 − CD103 + DCs, and XCR1 + CD103 − non-DCs, respectively (mean ± SD, n = 3 rats each, *P < 0.05).

Antigen-Labeled Donor T Cells Can Induce Specific Antibody Production

The highly efficient function of T cells in delivering alloantigen to the resident DCs in SLOs and inducing antibody production prompted us to harness donor T cells with antigens as a model of vaccine vectors for prophylactic antibody production. We labeled T cells with either fluorescein isothiocyanate (FITC) or R-phycoerythrin (PE) as a preliminary study. Recipient spleens after injection of labeled allogeneic T cells or soluble FITC-keyhole limpet hemocyanin (KLH) as a control contained specific anti-FITC antibody-forming cells in the outer margin of the PALS, ( Figure 8A ) and recipient sera contained anti-FITC antibodies (data not shown). When proliferation of FITC-labeled T cells was inhibited by MMC ( Figures 8B,C ) and i.v. injected into splenectomized rats, they readily migrated to the LNs ( Figure 8D ). Allogeneic, but not syngeneic, T cells induced anti-FITC antibodies, though their titers were low ( Figure 8E ).

Antigen-labeled donor T cells induce specific antibody production. (A) Three-color immunostaining of recipient spleen for anti-FITC antibody (blue), type IV collagen (brown), and BrdU (red). FITC-T cells induced specific anti-FITC antibody forming cell response (blue) in the outer margin of the PALS (O) at 7 days after injection [FITC-T]. When FITC-streptavidin was omitted for background staining, anti-FITC antibody was not detected [FITC-T BG]. Positive control staining for anti-FITC antibody forming cells 5 days after i.v. injection of FITC-labeled KLH [FITC-KLH]. G, germinal center P, PALS. Scale bar = 100 μm. (B) Experimental protocol for inducing antibody production in LNs with FITC-labeled donor T cells. (C) Effect of mitomycin C (MMC) on FITC-labeled donor T cell proliferation. Purity of FITC-labeled donor T cells was 94% by FCM. The proliferation of MMC-treated T cells induced by CD28 superagonist (SAg) was abrogated compared to intense proliferation in PBS-treated control. (D) Two-color immunostaining for donor MHCI (blue) and type IV collagen (brown) in the LNs 1 d after injection of MMC-treated FITC-labeled donor T cells. They readily migrated to the T cell area of the LNs (paracortex, PC). F, lymphoid follicle HEV, high endothelial venule. Scale bar = 100 μm. (E) MMC-treated FITC-labeled allogeneic, but not syngeneic, T cells induced a low, but significant, antibody response to FITC. (C,E) mean ± SD, n = 3 rats each, *P < 0.05. MFI, mean fluorescent intensity.

Specific Antibody Production Without DST Antibodies in F1 Rats That Received Parental Antigen-Labeled T Cells

In order to enhance the antibody response to labeled antigens and to avoid unnecessary alloantibody production, we attempted to inhibit the DST antibody response. We employed the semiallogeneic parental ACI donor and (Lewis × ACI)F1 hybrid recipient combination in which recipient T cells (RT1.A al ) share parental MHCI antigens (RT1.A a ) and cannot undergo the DST response. In this setting, recipient NK cells could kill parental cells via Ly49c, known as hybrid resistance (35). When MMC-treated and antigen-labeled parental T cells were injected into F1 recipients just after splenectomy ( Figure 9A ), they were readily phagocytosed by XCR1 + DCs in the LNs (data not shown) and 7 day sera had considerably higher titers of antibody than the allogeneic recipient group ( Figures 9B,D ). As expected, DST (anti-donor RT1.A a ) antibodies were negligible in the semiallogeneic F1 recipients, though an intense DST antibody response was induced in the allogeneic recipients ( Figures 9B,D ). The control group that received equivalent amounts of free PE showed the low level of anti-PE antibodies (Figure S7). Immunohistology of the LNs revealed the polytopical presence of specific anti-FITC or anti-PE antibody-forming cells ( Figures 9C,E ).

Antibody production to labeled antigens without DST antibodies in F1 rats that received parental T cells. (A) Experimental protocol for detecting specific antibody production in sera and the LNs. T cells were labeled either with FITC directly or with PE-conjugated ant-rat CD4 mAb. AFC, antibody-forming cells. (B,D) Antibody responses in parental ACI (RT1.A a B a ) to (Lewis × ACI)F1 hybrid rat (RT1.A al B al ) combination concerning anti-donor MHCI, anti-FITC (B), and anti-PE (D) antibodies in recipient sera. Both anti-FITC and anti-PE antibodies of parental to F1 rat combination had considerably higher titers than those of the allogeneic control. In contrast, DST (anti-RT1.A a ) antibodies were negligible, whereas, allogeneic control induced an intense level of DST antibodies (mean ± SD, n = 3 rats each, *P < 0.05). MFI, mean fluorescent intensity. (C,E) Double immunostaining for anti-FITC (blue, C) or anti-PE (blue, D) and type IV collagen (brown) in F1 rat LNs 7 days after antigen-labeled parental T cell injection. Specific antibody-forming cells (AFCs, blue) against labeled antigen were detected polytopically in the medullary cord of F1 rat LNs 7 days after injection. C, cervical (upper panel) and mesenteric (lower panel) LNs E, mediastinal LNs MC, medullary cord MS, medullary sinus. Scale bar = 100 μm (C,E) or 20 μm (C,E insets).


Red cell transfusion Edit

Historically, red blood cell transfusion was considered when the hemoglobin level fell below 10 g/dL or hematocrit fell below 30%. [2] [3] Because each unit of blood given carries risks, a trigger level lower than that, at 7 to 8 g/dL, is now usually used, as it has been shown to have better patient outcomes. [4] [5] The administration of a single unit of blood is the standard for hospitalized people who are not bleeding, with this treatment followed with re-assessment and consideration of symptoms and hemoglobin concentration. [4] Patients with poor oxygen saturation may need more blood. [4] The advisory caution to use blood transfusion only with more severe anemia is in part due to evidence that outcomes are worsened if larger amounts are given. [6] One may consider transfusion for people with symptoms of cardiovascular disease such as chest pain or shortness of breath. [3] In cases where patients have low levels of hemoglobin due to iron deficiency, but are cardiovascularly stable, parenteral iron is a preferred option based on both efficacy and safety. [7] Other blood products are given where appropriate, e.g., to treat clotting deficiencies.

Before a blood transfusion is given, there are many steps taken to ensure quality of the blood products, compatibility, and safety to the recipient. In 2012, a national blood policy was in place in 70% of countries and 69% of countries had specific legislation that covers the safety and quality of blood transfusion. [8]

Blood donation Edit

Blood transfusions use as sources of blood either one's own (autologous transfusion), or someone else's (allogeneic or homologous transfusion). The latter is much more common than the former. Using another's blood must first start with donation of blood. Blood is most commonly donated as whole blood obtained intravenously and mixed with an anticoagulant. In developed countries, donations are usually anonymous to the recipient, but products in a blood bank are always individually traceable through the whole cycle of donation, testing, separation into components, storage, and administration to the recipient. This enables management and investigation of any suspected transfusion related disease transmission or transfusion reaction. In developing countries, the donor is sometimes specifically recruited by or for the recipient, typically a family member, and the donation occurs immediately before the transfusion.

It is unclear whether applying alcohol swab alone or alcohol swab followed by antiseptic is able to reduce contamination of donor's blood. [9]

Processing and testing Edit

Donated blood is usually subjected to processing after it is collected, to make it suitable for use in specific patient populations. Collected blood is then separated into blood components by centrifugation: red blood cells, plasma, platelets, albumin protein, clotting factor concentrates, cryoprecipitate, fibrinogen concentrate, and immunoglobulins (antibodies). Red cells, plasma and platelets can also be donated individually via a more complex process called apheresis.

  • The World Health Organization (WHO) recommends that all donated blood be tested for transfusion transmissible infections. These include HIV, Hepatitis B, Hepatitis C, Treponema pallidum (syphilis) and, where relevant, other infections that pose a risk to the safety of the blood supply, such as Trypanosoma cruzi (Chagas disease) and Plasmodium species (malaria). [10] According to the WHO, 25 countries are not able to screen all donated blood for one or more of: HIV Hepatitis B Hepatitis C or syphilis. [11] One of the main reasons for this is because testing kits are not always available. [11] However the prevalence of transfusion-transmitted infections is much higher in low income countries compared to middle and high income countries. [11]
  • All donated blood should also be tested for the ABO blood group system and Rh blood group system to ensure that the patient is receiving compatible blood. [12]
  • In addition, in some countries platelet products are also tested for bacterial infections due to its higher inclination for contamination due to storage at room temperature. [13][14] Presence of cytomegalovirus (CMV) may also be tested because of the risk to certain immunocompromised recipients if given, such as those with organ transplant or HIV. However, not all blood is tested for CMV because only a certain amount of CMV-negative blood needs to be available to supply patient needs. Other than positivity for CMV, any products tested positive for infections are not used. [15]
  • Leukocyte reduction is the removal of white blood cells by filtration. Leukoreduced blood products are less likely to cause HLA alloimmunization (development of antibodies against specific blood types), febrile non-hemolytic transfusion reaction, cytomegalovirus infection, and platelet-transfusion refractoriness. [16]
  • Pathogen Reduction treatment that involves, for example, the addition of riboflavin with subsequent exposure to UV light has been shown to be effective in inactivating pathogens (viruses, bacteria, parasites and white blood cells) in blood products. [17][18][19] By inactivating white blood cells in donated blood products, riboflavin and UV light treatment can also replace gamma-irradiation as a method to prevent graft-versus-host disease (TA-GvHD). [20][21][22]

Compatibility testing Edit

Before a recipient receives a transfusion, compatibility testing between donor and recipient blood must be done. The first step before a transfusion is given is to type and screen the recipient's blood. Typing of recipient's blood determines the ABO and Rh status. The sample is then screened for any alloantibodies that may react with donor blood. [23] It takes about 45 minutes to complete (depending on the method used). The blood bank scientist also checks for special requirements of the patient (e.g. need for washed, irradiated or CMV negative blood) and the history of the patient to see if they have previously identified antibodies and any other serological anomalies.

A positive screen warrants an antibody panel/investigation to determine if it is clinically significant. An antibody panel consists of commercially prepared group O red cell suspensions from donors that have been phenotyped for antigens that correspond to commonly encountered and clinically significant alloantibodies. Donor cells may have homozygous (e.g. K+k+), heterozygous (K+k-) expression or no expression of various antigens (K−k−). The phenotypes of all the donor cells being tested are shown in a chart. The patient's serum is tested against the various donor cells. Based on the reactions of the patient's serum against the donor cells, a pattern will emerge to confirm the presence of one or more antibodies. Not all antibodies are clinically significant (i.e. cause transfusion reactions, HDN, etc.). Once the patient has developed a clinically significant antibody it is vital that the patient receive antigen-negative red blood cells to prevent future transfusion reactions. A direct antiglobulin test (Coombs test) is also performed as part of the antibody investigation. [24]

If there is no antibody present, an immediate spin crossmatch or computer-assisted crossmatch is performed where the recipient serum and donor rbc are incubated. In the immediate spin method, two drops of patient serum are tested against a drop of 3–5% suspension of donor cells in a test tube and spun in a serofuge. Agglutination or hemolysis (i.e., positive Coombs test) in the test tube is a positive reaction and the unit should not be transfused.

If an antibody is suspected, potential donor units must first be screened for the corresponding antigen by phenotyping them. Antigen negative units are then tested against the patient plasma using an antiglobulin/indirect crossmatch technique at 37 degrees Celsius to enhance reactivity and make the test easier to read.

In urgent cases where crossmatching cannot be completed, and the risk of dropping hemoglobin outweighs the risk of transfusing uncrossmatched blood, O-negative blood is used, followed by crossmatch as soon as possible. O-negative is also used for children and women of childbearing age. It is preferable for the laboratory to obtain a pre-transfusion sample in these cases so a type and screen can be performed to determine the actual blood group of the patient and to check for alloantibodies.

Compatibility of ABO and Rh system for Red Cell (Erythrocyte) Transfusion Edit

This chart shows possible matches in blood transfusion between donor and receiver using ABO and Rh system.

O- O+ B- B+ A- A+ AB- AB+
Recipient AB+

In the same way that the safety of pharmaceutical products is overseen by pharmacovigilance, the safety of blood and blood products is overseen by haemovigilance. This is defined by the World Health Organization (WHO) as a system ". to identify and prevent occurrence or recurrence of transfusion related unwanted events, to increase the safety, efficacy and efficiency of blood transfusion, covering all activities of the transfusion chain from donor to recipient." The system should include monitoring, identification, reporting, investigation and analysis of adverse events near-misses and reactions related to transfusion and manufacturing. [25] In the UK this data is collected by an independent organisation called SHOT (Serious Hazards Of Transfusion). [26]

Transfusions of blood products are associated with several complications, many of which can be grouped as immunological or infectious. There is controversy on potential quality degradation during storage. [27]

Immunologic reaction Edit

  • Acute hemolytic reactions are defined according to Serious Hazards of Transfusion (SHOT) as "fever and other symptoms/signs of haemolysis within 24 hours of transfusion confirmed by one or more of the following: a fall of Hb, rise in lactate dehydrogenase (LDH), positive direct antiglobulin test (DAT), positive crossmatch" [28] This is due to destruction of donor red blood cells by preformed recipient antibodies. Most often this occurs because of clerical errors or improper ABO blood typing and crossmatching resulting in a mismatch in ABO blood type between the donor and the recipient. Symptoms include fever, chills, chest pain, back pain, [29] hemorrhage, increased heart rate, shortness of breath, and rapid drop in blood pressure. When suspected, transfusion should be stopped immediately, and blood sent for tests to evaluate for presence of hemolysis. Treatment is supportive. Kidney injury may occur because of the effects of the hemolytic reaction (pigment nephropathy). [30] The severity of the transfusion reaction is depended upon amount of donor's antigen transfused, nature of the donor's antigens, the nature and the amount of recipient antibodies. [29]
  • Delayed hemolytic reactions occur more than 24 hours after a transfusion. They usually occur within 28 days of a transfusion. They can be due to either a low level of antibodies present prior to the start of the transfusion, which are not detectable on pre-transfusion testing or development of a new antibody against an antigen in the transfused blood. Therefore, delayed haemolytic reaction does not manifest until after 24 hours when enough amount of antibodies are available to cause a reaction. The red blood cells are removed by macrophages from the blood circulation into liver and spleen to be destroyed, which leads to extravascular haemolysis. This process usually mediated by anti-Rh and anti-Kidd antibodies. However, this type of transfusion reaction is less severe when compared to acute haemolytic transfusion reaction. [29]
  • Febrile nonhemolytic reactions are, along with allergic transfusion reactions, the most common type of blood transfusion reaction and occur because of the release of inflammatory chemical signals released by white blood cells in stored donor blood [16] or attack on donor's white blood cells by recipient's antibodies. [29] This type of reaction occurs in about 7% of transfusions. Fever is generally short lived and is treated with antipyretics, and transfusions may be finished as long as an acute hemolytic reaction is excluded. This is a reason for the now-widespread use of leukoreduction – the filtration of donor white cells from red cell product units. [16]
  • Allergic transfusion reactions are caused by IgE anti-allergen antibodies. When antibodies are bound to its antigens, histamine is released from mast cells and basophils. Either IgE antibodies from the donor's or recipient's side can cause the allergic reaction. It is more common in patients who have allergic conditions such as hay fever. Patient may feel itchy or having hives but the symptoms are usually mild and can be controlled by stopping the transfusion and giving antihistamines. [29]
  • Anaphylactic reactions are rare life-threatening allergic conditions caused by IgA anti-plasma protein antibodies. For patients who have selective immunoglobulin A deficiency, the reaction is presumed to be caused by IgA antibodies in the donor's plasma. The patient may present with symptoms of fever, wheezing, coughing, shortness of breath, and circulatory shock. Urgent treatment with epinephrine is needed. [29]
  • Post-transfusion purpura is an extremely rare complication that occurs after blood product transfusion and is associated with the presence of antibodies in the patient's blood directed against both the donor's and recipient's platelets HPA (human platelet antigen). Recipients who lack this protein develop sensitization to this protein from prior transfusions or previous pregnancies, can develop thrombocytopenia, bleeding into the skin, and can display purplish discolouration of skin which is known as purpura. Intravenous immunoglobulin (IVIG) is treatment of choice. [29][31]
  • Transfusion-related acute lung injury (TRALI) is a syndrome that is similar to acute respiratory distress syndrome (ARDS), which develops during or within 6 hours of transfusion of a plasma-containing blood product. Fever, hypotension, shortness of breath, and tachycardia often occurs in this type of reaction. For a definitive diagnosis to be made, symptoms must occur within 6 hours of transfusion, hypoxemia must be present, there must be radiographic evidence of bilateral infiltrates and there must be no evidence of left atrial hypertension (fluid overload). [32] It occurs in 15% of the transfused patient with mortality rate of 5 to 10%. Recipient risk factors includes: end-stage liver disease, sepsis, haematological malignancies, sepsis, and ventilated patients. Antibodies to human neutrophil antigens (HNA) and human leukocyte antigens (HLA) have been associated with this type of transfusion reaction. Donor's antibodies interacting with antigen positive recipient tissue result in release of inflammatory cytokines, resulting in pulmonary capillary leakage. The treatment is supportive. [33] is a common, yet underdiagnosed, reaction to blood product transfusion consisting of the new onset or exacerbation of three of the following within 6 hours of cessation of transfusion: acute respiratory distress, elevated brain natriuretic peptide (BNP), elevated central venous pressure (CVP), evidence of left heart failure, evidence of positive fluid balance, and/or radiographic evidence of pulmonary edema. [32]
  • Transfusion-associated graft versus host disease frequently occurs in immunodeficient patients where recipient's body failed to eliminate donor's T cells. Instead, donor's T cells attack the recipient's cells. It occurs one week after transfusion. [29] Fever, rash, diarrhoea are often associated with this type of transfusion reaction. Mortality rate is high, with 89.7% of the patients dead after 24 days. Immunosuppressive treatment is the most common way of treatment. [34] Irradiation and leukoreduction of blood products is necessary for high risk patients for prevent T cells from attacking recipient cells. [29]

Infection Edit

The use of greater amount of red blood cells is associated with a high risk of infections. In those who were given red blood only with significant anemia infection rates were 12% while in those who were given red blood at milder levels of anemia infection rates were 17%. [35] [ clarification needed ]

On rare occasions, blood products are contaminated with bacteria. This can result in a life-threatening infection known as transfusion-transmitted bacterial infection. The risk of severe bacterial infection is estimated, as of 2002 [update] , at about 1 in 50,000 platelet transfusions, and 1 in 500,000 red blood cell transfusions. [36] Blood product contamination, while rare, is still more common than actual infection. The reason platelets are more often contaminated than other blood products is that they are stored at room temperature for short periods of time. Contamination is also more common with longer duration of storage, especially if that means more than 5 days. Sources of contaminants include the donor's blood, donor's skin, phlebotomist's skin, and containers. Contaminating organisms vary greatly, and include skin flora, gut flora, and environmental organisms. There are many strategies in place at blood donation centers and laboratories to reduce the risk of contamination. A definite diagnosis of transfusion-transmitted bacterial infection includes the identification of a positive culture in the recipient (without an alternative diagnosis) as well as the identification of the same organism in the donor blood.

Since the advent of HIV testing of donor blood in the mid/later 1980s, ex. 1985's ELISA, the transmission of HIV during transfusion has dropped dramatically. Prior testing of donor blood only included testing for antibodies to HIV. However, because of latent infection (the "window period" in which an individual is infectious, but has not had time to develop antibodies) many cases of HIV seropositive blood were missed. The development of a nucleic acid test for the HIV-1 RNA has dramatically lowered the rate of donor blood seropositivity to about 1 in 3 million units. As transmittance of HIV does not necessarily mean HIV infection, the latter could still occur at an even lower rate.

The transmission of hepatitis C via transfusion currently stands at a rate of about 1 in 2 million units. As with HIV, this low rate has been attributed to the ability to screen for both antibodies as well as viral RNA nucleic acid testing in donor blood.

Other rare transmissible infections include hepatitis B, syphilis, Chagas disease, cytomegalovirus infections (in immunocompromised recipients), HTLV, and Babesia.

Comparison table Edit

Comparison of symptoms of blood transfusion reactions characterized by fever. [37]
+ =Occasionally present ++ =Frequently present
Febrile nonhemolytic TRALI Acute hemolytic Bacterial contamination
Appearance of symptoms during or after transfusion Usually toward end. 5-10% appear up to 2 hours after. Early (after 10-15 ml) Early (after 50-100 ml) Up to 8 hours after transfusion
Fever + ++ ++ ++
Chills ++ ++ ++ +++
Cold ++ - + -
Discomfort ++ - - -
Rigors + - - -
Headache + - + -
Nausea and/or vomiting + - ++ -
Dyspnea + ++ ++ -
Cyanosis - ++ ++ -
Hypotension / circulatory shock - ++ ++ ++
Disseminated intravascular coagulation - - ++ ++
Hemoglobinuria - - ++ +
Renal failure - - ++ ++
Back pain - - ++ -

Inefficacy Edit

Transfusion inefficacy or insufficient efficacy of a given unit(s) of blood product, while not itself a "complication" per se, can nonetheless indirectly lead to complications – in addition to causing a transfusion to fully or partly fail to achieve its clinical purpose. This can be especially significant for certain patient groups such as critical-care or neonatals.

For red blood cells (RBC), by far the most commonly transfused product, poor transfusion efficacy can result from units damaged by the so-called storage lesion – a range of biochemical and biomechanical changes that occur during storage. With red cells, this can decrease viability and ability for tissue oxygenation. [38] Although some of the biochemical changes are reversible after the blood is transfused, [39] the biomechanical changes are less so, [40] and rejuvenation products are not yet able to adequately reverse this phenomenon. [41] There has been controversy about whether a given product unit's age is a factor in transfusion efficacy, specifically about whether "older" blood directly or indirectly increases risks of complications. [42] [43] Studies have not been consistent on answering this question, [44] with some showing that older blood is indeed less effective but with others showing no such difference these developments are being closely followed by hospital blood bankers – who are the physicians, typically pathologists, who collect and manage inventories of transfusable blood units.

Certain regulatory measures are in place to minimize RBC storage lesion – including a maximum shelf life (currently 42 days), a maximum auto-hemolysis threshold (currently 1% in the US, 0.8% in Europe), and a minimum level of post-transfusion RBC survival in vivo (currently 75% after 24 hours). [45] However, all of these criteria are applied in a universal manner that does not account for differences among units of product. [46] For example, testing for the post-transfusion RBC survival in vivo is done on a sample of healthy volunteers, and then compliance is presumed for all RBC units based on universal (GMP) processing standards (of course, RBC survival by itself does not guarantee efficacy, but it is a necessary prerequisite for cell function, and hence serves as a regulatory proxy). Opinions vary as to the "best" way to determine transfusion efficacy in a patient in vivo. [47] In general, there are not yet any in vitro tests to assess quality or predict efficacy for specific units of RBC blood product prior to their transfusion, though there is exploration of potentially relevant tests based on RBC membrane properties such as erythrocyte deformability [48] and erythrocyte fragility (mechanical). [49]

Physicians have adopted a so-called "restrictive protocol" – whereby transfusion is held to a minimum – in part because of the noted uncertainties surrounding storage lesion, in addition to the very high direct and indirect costs of transfusions. [50] [51] [52] Of course, restrictive protocol is not an option with some especially vulnerable patients who may require the best possible efforts to rapidly restore tissue oxygenation.

Although transfusions of platelets are far less numerous (relative to RBC), platelet storage lesion and resulting efficacy loss is also a concern. [53]

Other Edit

  • A known relationship between intra-operative blood transfusion and cancer recurrence has been established in colorectal cancer. [54] In lung cancer intra-operative blood transfusion has been associated with earlier recurrence of cancer, worse survival rates and poorer outcomes after lung resection. [55][56] Also studies shown to us [who?] , failure of the immune system caused by blood transfusion can be categorized as one of the main factors leading to more than 10 different cancer types that are fully associated with blood transfusion and the innate and adaptive immune system. [57] Allogeneic blood transfusion, through five major mechanisms including the lymphocyte-T set, myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), natural killer cells (NKCs), and dendritic cells (DCs) can help the recipient's defense mechanisms. On the other hand, the role for each of the listed items includes activation of the antitumorCD8+cytotoxic T lymphocytes (CD8+/CTL), temporal inactivation of Tregs, inactivation of the STAT3 signaling pathway, the use of bacteria to enhance the antitumor immune response and cellular Immunotherapy. [58]
  • Transfusion-associated volume overload is a common complication simply because blood products have a certain amount of volume. This is especially the case in recipients with underlying cardiac or kidney disease. Red cell transfusions can lead to volume overload when they must be repeated because of insufficient efficacy (see above). Plasma transfusion is especially prone to causing volume overload because of its hypertonicity.
  • It has been proved that blood transfusion produce worse outcomes after cytoreductive surgery and HIPEC. [59]
  • Hypothermia can occur with transfusions with large quantities of blood products which normally are stored at cold temperatures. Core body temperature can go down as low as 32 °C and can produce physiologic disturbances. Prevention should be done with warming the blood to ambient temperature prior to transfusions.
  • Transfusions with large amounts of red blood cells, whether due to severe hemorrhaging and/or transfusion inefficacy (see above), can lead to an inclination for bleeding. The mechanism is thought to be due to disseminated intravascular coagulation, along with dilution of recipient platelets and coagulation factors. Close monitoring and transfusions with platelets and plasma is indicated when necessary.
  • Metabolic alkalosis can occur with massive blood transfusions because of the breakdown of citrate stored in blood into bicarbonate.
  • Hypocalcemia can also occur with massive blood transfusions because of the complex of citrate with serum calcium. Calcium levels below 0.9 mmol/L should be treated. [60] is often used by athletes, drug addicts or military personnel for reasons such as to increase physical stamina, to fake a drug detection test or simply to remain active and alert during the duty-times respectively. However a lack of knowledge and insufficient experience can turn a blood transfusion into a sudden death. For example, when individuals run the frozen blood sample directly in their veins this cold blood rapidly reaches the heart, where it disturbs the heart's original pace leading to cardiac arrest and sudden death.

Globally around 85 million units of red blood cells are transfused in a given year. [3]

In the United States, blood transfusions were performed nearly 3 million times during hospitalizations in 2011, making it the most common procedure performed. The rate of hospitalizations with a blood transfusion nearly doubled from 1997, from a rate of 40 stays to 95 stays per 10,000 population. It was the most common procedure performed for patients 45 years of age and older in 2011, and among the top five most common for patients between the ages of 1 and 44 years. [61]

According to the New York Times: "Changes in medicine have eliminated the need for millions of blood transfusions, which is good news for patients getting procedures like coronary bypasses and other procedures that once required a lot of blood." And, "Blood bank revenue is falling, and the decline may reach $1.5 billion a year this year [2014] from a high of $5 billion in 2008." Job losses will reach as high as 12,000 within the next three to five years, roughly a quarter of the total in the industry, according to the Red Cross. [62]

Beginning with William Harvey's experiments on the circulation of blood, recorded research into blood transfusion began in the 17th century, with successful experiments in transfusion between animals. However, successive attempts by physicians to transfuse animal blood into humans gave variable, often fatal, results. [63]

Pope Innocent VIII is sometimes said to have been given "the world's first blood transfusion" by his physician Giacomo di San Genesio, who had him drink (by mouth) the blood of three 10-year-old boys. The boys subsequently died. The evidence for this story, however, is unreliable and considered a possible anti-Jewish blood libel. [64]

Early attempts Edit

The first reported successful blood transfusions were performed by the Incas as early as the 1500s. [65] Spanish conquistadors witnessed blood transfusions when they arrived in the sixteenth century. [66] The prevalence of type O blood among Indigenous people of the Andean region meant such procedures would have held less risk than blood transfusion attempts among populations with incompatible blood types, which contributed to the failures of early attempts in Europe. [66]

Animal blood Edit

Working at the Royal Society in the 1660s, the physician Richard Lower began examining the effects of changes in blood volume on circulatory function and developed methods for cross-circulatory study in animals, obviating clotting by closed arteriovenous connections. The new instruments he was able to devise enabled him to perform the first reliably documented successful transfusion of blood in front of his distinguished colleagues from the Royal Society.

According to Lower's account, ". towards the end of February 1665 [I] selected one dog of medium size, opened its jugular vein, and drew off blood, until its strength was nearly gone. Then, to make up for the great loss of this dog by the blood of a second, I introduced blood from the cervical artery of a fairly large mastiff, which had been fastened alongside the first, until this latter animal showed … it was overfilled … by the inflowing blood." After he "sewed up the jugular veins", the animal recovered "with no sign of discomfort or of displeasure".

Lower had performed the first blood transfusion between animals. He was then "requested by the Honorable [Robert] Boyle … to acquaint the Royal Society with the procedure for the whole experiment", which he did in December 1665 in the Society's Philosophical Transactions. [67]

The first blood transfusion from animal to human was administered by Dr. Jean-Baptiste Denys, eminent physician to King Louis XIV of France, on June 15, 1667. [68] He transfused the blood of a sheep into a 15-year-old boy, who survived the transfusion. [69] Denys performed another transfusion into a labourer, who also survived. Both instances were likely due to the small amount of blood that was actually transfused into these people. This allowed them to withstand the allergic reaction.

Denys's third patient to undergo a blood transfusion was Swedish Baron Gustaf Bonde. He received two transfusions. After the second transfusion Bonde died. [70] In the winter of 1667, Denys performed several transfusions on Antoine Mauroy with calf's blood. On the third account Mauroy died. [71]

Six months later in London, Lower performed the first human transfusion of animal blood in Britain, where he "superintended the introduction in [a patient's] arm at various times of some ounces of sheep's blood at a meeting of the Royal Society, and without any inconvenience to him." The recipient was Arthur Coga, "the subject of a harmless form of insanity." Sheep's blood was used because of speculation about the value of blood exchange between species it had been suggested that blood from a gentle lamb might quiet the tempestuous spirit of an agitated person and that the shy might be made outgoing by blood from more sociable creatures. Coga received 20 shillings (equivalent to £173 in 2019) to participate in the experiment. [72]

Lower went on to pioneer new devices for the precise control of blood flow and the transfusion of blood his designs were substantially the same as modern syringes and catheters. [67] Shortly after, Lower moved to London, where his growing practice soon led him to abandon research. [73]

These early experiments with animal blood provoked a heated controversy in Britain and France. [70] Finally, in 1668, the Royal Society and the French government both banned the procedure. The Vatican condemned these experiments in 1670. Blood transfusions fell into obscurity for the next 150 years. [ citation needed ]

Human blood Edit

The science of blood transfusion dates to the first decade of the 20th century, with the discovery of distinct blood types leading to the practice of mixing some blood from the donor and the receiver before the transfusion (an early form of cross-matching).

In the early 19th century, British obstetrician Dr. James Blundell made efforts to treat hemorrhage by transfusion of human blood using a syringe. In 1818 following experiments with animals, he performed the first successful transfusion of human blood to treat postpartum hemorrhage. Blundell used the patient's husband as a donor, and extracted four ounces of blood from his arm to transfuse into his wife. During the years 1825 and 1830, Blundell performed 10 transfusions, five of which were beneficial, and published his results. He also invented a number of instruments for the transfusion of blood. [74] He made a substantial amount of money from this endeavour, roughly $2 million ($50 million real dollars). [75]

In 1840, at St George's Hospital Medical School in London, Samuel Armstrong Lane, aided by Dr. Blundell, performed the first successful whole blood transfusion to treat haemophilia.

However, early transfusions were risky and many resulted in the death of the patient. By the late 19th century, blood transfusion was regarded as a risky and dubious procedure, and was largely shunned by the medical establishment.

Work to emulate James Blundell continued in Edinburgh. In 1845 the Edinburgh Journal described the successful transfusion of blood to a woman with severe uterine bleeding. Subsequent transfusions were successful with patients of Professor James Young Simpson after whom the Simpson Memorial Maternity Pavilion in Edinburgh was named. [76]

Various isolated reports of successful transfusions emerged towards the end of the 19th century. [77] The largest series of early successful transfusions took place at the Edinburgh Royal Infirmary between 1885 and 1892. Edinburgh later became the home of the first blood donation and blood transfusion services. [76]

20th century Edit

Only in 1901, when the Austrian Karl Landsteiner discovered three human blood groups (O, A, and B), did blood transfusion achieve a scientific basis and became safer.

Landsteiner discovered that adverse effects arise from mixing blood from two incompatible individuals. He found that mixing incompatible types triggers an immune response and the red blood-cells clump. The immunological reaction occurs when the receiver of a blood transfusion has antibodies against the donor blood-cells. The destruction of red blood cells releases free hemoglobin into the bloodstream, which can have fatal consequences. Landsteiner's work made it possible to determine blood group and allowed blood transfusions to take place much more safely. For his discovery he won the Nobel Prize in Physiology and Medicine in 1930 many other blood groups have been discovered since.

George Washington Crile is credited with performing the first surgery using a direct blood transfusion in 1906 at St. Alexis Hospital in Cleveland while a professor of surgery at Case Western Reserve University. [78]

Jan Janský also discovered the human blood groups in 1907 he classified blood into four groups: I, II, III, IV. [79] His nomenclature is still used in Russia and in states of the former USSR, in which blood types O, A, B, and AB are respectively designated I, II, III, and IV.

Dr. William Lorenzo Moss's (1876–1957) Moss-blood typing technique of 1910 was widely used until World War II. [80] [81]

William Stewart Halsted, M.D. (September 23, 1852 – September 7, 1922), an American surgeon, performed one of the first blood transfusions in the United States. He had been called to see his sister after she had given birth. He found her moribund from blood loss, and in a bold move withdrew his own blood, transfused his blood into his sister, and then operated on her to save her life.

Blood banks in WWI Edit

While the first transfusions had to be made directly from donor to receiver before coagulation, it was discovered that by adding anticoagulant and refrigerating the blood it was possible to store it for some days, thus opening the way for the development of blood banks. John Braxton Hicks was the first to experiment with chemical methods to prevent the coagulation of blood at St Mary's Hospital, London in the late-19th century. His attempts, using phosphate of soda, however, proved unsuccessful.

The Belgian doctor Albert Hustin performed the first non-direct transfusion on March 27, 1914, though this involved a diluted solution of blood. The Argentine doctor Luis Agote used a much less diluted solution in November of the same year. Both used sodium citrate as an anticoagulant. [82]

The First World War (1914-1918) acted as a catalyst for the rapid development of blood banks and transfusion techniques. Canadian doctor and Lieutenant Lawrence Bruce Robertson became instrumental in persuading the Royal Army Medical Corps to adopt the use of blood transfusion at the Casualty Clearing Stations for the wounded. In October 1915 Robertson performed his first wartime transfusion with a syringe to a patient suffering from multiple shrapnel wounds. He followed this up with four subsequent transfusions in the following months, and his success was reported to Sir Walter Morley Fletcher, director of the Medical Research Committee. [83]

Robertson published his findings in the British Medical Journal in 1916 and, with the help of a few like-minded individuals (including the eminent physician Edward William Archibald (1872-1945), who introduced the citrate anticoagulant method), was able to persuade the British authorities of the merits of blood transfusion. Robertson went on to establish the first blood-transfusion apparatus at a Casualty Clearing Station on the Western Front in the spring of 1917. [83] [84]

Oswald Hope Robertson, a medical researcher and U.S. Army officer, was attached to the RAMC in 1917, where he became instrumental in establishing the first blood banks in preparation for the anticipated Third Battle of Ypres. [85] He used sodium citrate as the anticoagulant blood was extracted from punctures in the vein and was stored in bottles at British and American Casualty Clearing Stations along the Front. Robertson also experimented with preserving separated red blood cells in iced bottles. [84] Geoffrey Keynes, a British surgeon, developed a portable machine that could store blood to enable transfusions to be carried out more easily.

Expansion Edit

The secretary of the British Red Cross, Percy Oliver, established the world's first blood-donor service in 1921. In that year, Oliver was contacted by King's College Hospital, where they were in urgent need of a blood donor. [86] After providing a donor, Oliver set about organizing a system for the voluntary registration of blood donors at clinics around London, with Sir Geoffrey Keynes appointed as a medical adviser. Volunteers were subjected to a series of physical tests to establish their blood group. The London Blood Transfusion Service was free of charge and expanded rapidly in its first few years of operation. By 1925 it was providing services for almost 500 patients it was incorporated into the structure of the British Red Cross in 1926. Similar systems developed in other cities, including Sheffield, Manchester and Norwich, and the service's work began to attract international attention. France, Germany, Austria, Belgium, Australia and Japan established similar services. [87]

Alexander Bogdanov founded an academic institution devoted to the science of blood transfusion in Moscow in 1925. Bogdanov was motivated, at least in part, by a search for eternal youth, and remarked with satisfaction on the improvement of his eyesight, suspension of balding, and other positive symptoms after receiving 11 transfusions of whole blood. Bogdanov died in 1928 as a result of one of his experiments, when the blood of a student suffering from malaria and tuberculosis was given to him in a transfusion. [88] Following Bogdanov's lead, Vladimir Shamov and Sergei Yudin in the USSR pioneered the transfusion of cadaveric blood from recently deceased donors. Yudin performed such a transfusion successfully for the first time on March 23, 1930 and reported his first seven clinical transfusions with cadaveric blood at the Fourth Congress of Ukrainian Surgeons at Kharkiv in September. However, this method was never used widely, even in the Soviet Union.

Frederic Durán-Jordà established one of the earliest blood banks during the Spanish Civil War in 1936. Duran joined the Transfusion Service at the Barcelona Hospital at the start of the conflict, but the hospital was soon overwhelmed by the demand for blood and the paucity of available donors. With support from the Department of Health of the Spanish Republican Army, Duran established a blood bank for the use of wounded soldiers and civilians. The 300–400 mL of extracted blood was mixed with 10% citrate solution in a modified Duran Erlenmeyer flask. The blood was stored in a sterile glass enclosed under pressure at 2 °C. During 30 months of work, the Transfusion Service of Barcelona registered almost 30,000 donors, and processed 9,000 liters of blood. [89]

In 1937 Bernard Fantus, director of therapeutics at the Cook County Hospital in Chicago, established the first hospital blood-bank in the United States. In setting up a hospital laboratory that preserved, refrigerated and stored donor blood, Fantus originated the term "blood bank". Within a few years, hospital and community blood-banks were established across the United States. [90]

Frederic Durán-Jordà fled to Britain in 1938 and worked with Dr Janet Vaughan at the Royal Postgraduate Medical School at Hammersmith Hospital to establish a system of national blood banks in London. [91] With the outbreak of war appearing imminent in 1938, the War Office created the Army Blood Supply Depot (ABSD) in Bristol, headed by Lionel Whitby and in control of four large blood-depots around the country. British policy through the war was to supply military personnel with blood from centralized depots, in contrast to the approach taken by the Americans and Germans where troops at the front were bled to provide required blood. The British method proved more successful in adequately meeting all requirements, and over 700,000 donors were bled [ by whom? ] over the course of the war. This system evolved into the National Blood Transfusion Service established in 1946, the first national service to be implemented. [92]

Stories tell of Nazis in Eastern Europe during World War II using captive children as repeated involuntary blood-donors. [93]

Medical advances Edit

A blood-collection program was initiated [ by whom? ] in the US in 1940 and Edwin Cohn pioneered the process of blood fractionation. He worked out the techniques for isolating the serum albumin fraction of blood plasma, which is essential for maintaining the osmotic pressure in the blood vessels, preventing their collapse.

Gordon R. Ward, writing in the correspondence columns of the British Medical Journal, proposed the use of blood plasma as a substitute for whole blood and for transfusion purposes as early as 1918. At the onset of World War II, liquid plasma was used in Britain. A large project, known as "Blood for Britain" began in August 1940 to collect blood in New York City hospitals for the export of plasma to Britain. A dried plasma package was developed, [ by whom? ] which reduced breakage and made transportation, packaging, and storage much simpler. [94]

The resulting dried plasma package came in two tin cans containing 400 mL bottles. One bottle contained enough distilled water to reconstitute the dried plasma contained within the other bottle. In about three minutes, the plasma would be ready to use and could stay fresh for around four hours. [95] Dr. Charles R. Drew was appointed medical supervisor, and he was able to transform the test-tube methods into the first successful technique for mass production.

Another important breakthrough came in 1937–40 when Karl Landsteiner (1868-1943), Alex Wiener, Philip Levine, and R.E. Stetson discovered the Rhesus blood group system, which was found to be the cause of the majority of transfusion reactions up to that time. Three years later, the introduction by J.F. Loutit and Patrick L. Mollison of acid–citrate–dextrose (ACD) solution, which reduced the volume of anticoagulant, permitted transfusions of greater volumes of blood and allowed longer-term storage.

Carl Walter and W.P. Murphy Jr. introduced the plastic bag for blood collection in 1950. Replacing breakable glass bottles with durable plastic bags made from PVC allowed for the evolution of a collection system capable of safe and easy preparation of multiple blood components from a single unit of whole blood.

In the field of cancer surgery, the replacement of massive blood-loss became a major problem. The cardiac-arrest rate was high. In 1963 C. Paul Boyan and William S. Howland discovered that the temperature of the blood and the rate of infusion greatly affected survival rates, and introduced blood warming to surgery. [96] [97]

Further extending the shelf-life of stored blood up to 42 days was an anticoagulant preservative, CPDA-1, introduced in 1979, which increased the blood supply and facilitated resource-sharing among blood banks. [98] [99]

As of 2006 [update] about 15 million units of blood products were transfused per year in the United States. [100] By 2013 the number had declined to about 11 million units, because of the shift towards laparoscopic surgery and other surgical advances and studies that have shown that many transfusions were unnecessary. For example, the standard of care reduced the amount of blood transfused in one case from 750 to 200 ml. [62]

Neonate Edit

To ensure the safety of blood transfusion to pediatric patients, hospitals are taking additional precautions to avoid infection and prefer to use specially tested pediatric blood units that are guaranteed negative for Cytomegalovirus. Most guidelines recommend the provision of CMV-negative blood components and not simply leukoreduced components for newborns or low birthweight infants in whom the immune system is not fully developed. [101] These specific requirements place additional restrictions on blood donors who can donate for neonatal use.

Neonatal transfusions typically fall into one of two categories:

  • "Top-up" transfusions, to replace losses due to investigational losses and correction of anemia.
  • Exchange (or partial exchange) transfusions are done for removal of bilirubin, removal of antibodies and replacement of red cells (e.g., for anemia secondary to thalassemias and other hemoglobinopathies). [102]

Significant blood loss Edit

A massive transfusion protocol is used when significant blood loss is present such as in major trauma, when more than ten units of blood are needed. Packed red blood cells, fresh frozen plasma, and platelets are generally administered. [103] Typically higher ratios of fresh frozen plasma and platelets are given relative to packed red blood cells. [103]

Unknown blood type Edit

Because blood type O negative is compatible with anyone, it is often overused and in short supply. [104] According to the American Association of Blood Banks, the use of this blood should be restricted to persons with O negative blood, as nothing else is compatible with them, and women who might be pregnant and for whom it would be impossible to do blood group testing before giving them emergency treatment. [104] Whenever possible, the AABB recommends that O negative blood be conserved by using blood type testing to identify a less scarce alternative. [104]

Religious objections Edit

Although there are clinical situations where transfusion with red blood cells is the only clinically appropriate option, clinicians look at whether alternatives are feasible. This can be due to several reasons, such as patient safety, economic burden or scarcity of blood. Guidelines recommend blood transfusions should be reserved for patients with or at risk of cardiovascular instability due to the degree of their anaemia. [106] [107] In these cases parenteral iron is recommended.

Thus far, there are no available oxygen-carrying blood substitutes, which is the typical objective of a blood (RBC) transfusion however, there are widely available non-blood volume expanders for cases where only volume restoration is required. These are helping doctors and surgeons avoid the risks of disease transmission and immune suppression, address the chronic blood donor shortage, and address the concerns of Jehovah's Witnesses and others who have religious objections to receiving transfused blood.

A number of blood substitutes have been explored (and still are), but thus far they all suffer from many challenges. Most attempts to find a suitable alternative to blood thus far have concentrated on cell-free hemoglobin solutions. Blood substitutes could make transfusions more readily available in emergency medicine and in pre-hospital EMS care. If successful, such a blood substitute could save many lives, particularly in trauma where massive blood loss results. Hemopure, a hemoglobin-based therapy, is approved for use in South Africa.

Minor blood transfusions are used by a minority of nyaope drug addicts in South Africa to economically share the high the drug induces in a practice colloquially known as Bluetoothing, named after the wireless technology of the same name. [108]

Veterinarians also administer transfusions to other animals. Various species require different levels of testing to ensure a compatible match. For example, cats have 3 known blood types, cattle have 11, dogs have 13, pigs have 16, and horses have 34. However, in many species (especially horses and dogs), cross matching is not required before the first transfusion, as antibodies against non-self cell surface antigens are not expressed constitutively – i.e. the animal has to be sensitized before it will mount an immune response against the transfused blood.

The rare and experimental practice of inter-species blood transfusions is a form of xenograft.


The ongoing coronavirus disease 2019 (COVID-19) global pandemic is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 also known as 2019-nCoV or HCoV-19) ( Coronaviridae Study Group of the International Committee on Taxonomy of Viruses, 2020 Jiang etਊl., 2020a , 2020c ). The four main genera of coronaviruses are known as α, β, γ, and δ. SARS-CoV-2, together with SARS-CoV-1, identified in 2003, and Middle East respiratory syndrome coronavirus (MERS-CoV), identified in 2012, belong to the β-CoV genus ( Coronaviridae Study Group of the International Committee on Taxonomy of Viruses, 2020 Zhou etਊl., 2020 Zhu etਊl., 2020 ).

For each coronavirus particle, the viral genome is packed with nucleocapsid (N) proteins and surrounded by an envelope containing structural proteins. One of these structural proteins (spike [S] protein) trimerizes and mediates viral entry into host cells ( Li, 2016 ), and it is the major target for human neutralizing antibodies ( Jiang etਊl., 2020b Premkumar etਊl., 2020 Wu etਊl., 2020a ). It contains 1,273 amino acids with a large ectodomain (S-ECD), one transmembrane helix, and a small intracellular C terminus (Figure S1A). Two major domains within S-ECD have been identified as S1 head and S2 stalk regions, and the crucial receptor-binding domain (RBD) localizes to the S1 portion (Figure S1A).

Because binding of SARS-CoV-2 RBD to its human receptor, angiotensin-converting enzyme 2 (ACE2), is a critical initial step for virus entry into target cells, blocking this interaction with antibodies is likely a promising approach for both treatment and protection. This would be especially true for broadly neutralizing antibodies targeting conserved epitopes present in different coronaviruses, such as SARS-CoV-1 and the newly emerging SARS-CoV-2, both of which are from the same coronavirus subfamily and share the same human receptor ACE2.

Efforts to obtain human neutralizing antibodies against the S protein have involved a variety of methods as phage display ( Liu etਊl., 2020b Sun etਊl., 2020 Wu etਊl., 2020b ), humanized mice ( Hansen etਊl., 2020 ), antibody screening from SARS-CoV-1-recovered individuals ( Pinto etਊl., 2020 Wec etਊl., 2020 ), and single B cell antibody cloning from SARS-CoV-2 convalescent donors ( Andreano etਊl., 2020 Brouwer etਊl., 2020 Cao etਊl., 2020 Chen etਊl., 2020 Chi etਊl., 2020 Ju etਊl., 2020 Kreer etਊl., 2020 Liu etਊl., 2020a Robbiani etਊl., 2020 Rogers etਊl., 2020 Seydoux etਊl., 2020 Wan etਊl., 2020 Wu etਊl., 2020c Zost etਊl., 2020a , 2020b ). The neutralizing activities of these cloned antibodies are radically different, with 50% inhibitory concentration (IC50) values ranging from single-digit nanograms per milliliter (ng/mL) to non-neutralizing. The antibodies that bind to RBD showed generally higher neutralization potency compared with antibodies with non-RBD epitopes ( Rogers etਊl., 2020 ). Although some of the reported antibodies showed cross-neutralizing activity ( Lv etਊl., 2020 Pinto etਊl., 2020 Wec etਊl., 2020 ), their potency against SARS-CoV-1 and SARS-CoV-2 was not equally high. Moreover, to the best of our knowledge, the antibody-dependent enhancement (ADE) effect of these antibodies has never been evaluated, and accordingly, the relationship between ADE and different SARS-CoV-2 S protein epitopes has not been determined.

Antibody-bound virus particles could be attached on the surface of immune cells through Fcγ-receptor-mediated internalization for subsequent degradation. However, instead of protection, antibody binding might facilitate viral particles to enter and invade host cells. This � of viral entry” phenomenon has been documented for many viruses, including dengue, Zika, and SARS-CoV-1 viruses ( Eroshenko etਊl., 2020 Iwasaki and Yang, 2020 Katzelnick etਊl., 2017 Miner and Diamond, 2017 Salje etਊl., 2018 ). In SARS-CoV-1 infection, antibodies binding the S protein facilitate ACE2-independent virus internalization into macrophages, monocytes, and B cells in vitro ( Jaume etਊl., 2011 Wang etਊl., 2014 Yip etਊl., 2014 ). Nevertheless, viral uptake does not necessarily result in a productive viral infection, meaning that ADE of viral entry in vitro does not predict ADE of infection and ADE of disease ( Arvin etਊl., 2020 Halstead and Katzelnick, 2020 ). For example, viral replication was abortive in vitro despite enhancement of SARS-CoV-1 virus entry into a B cell line, Raji cells ( Jaume etਊl., 2011 ). Whether antibodies against SARS-CoV-2 could induce ADE of viral entry and whether the invaded viruses undergo active replication are both still unknown.

Here, we selected a convalescent individual with a high level of serum IgG neutralizing activity against SARS-CoV-2 and isolated many expanded clones of memory B cells expressing closely related antibodies with the same Ig variable gene segments and highly similar CDR3 sequences. Most of these isolated antibodies targeted four groups of five distinct epitopes on the RBD of S protein. Characterization of both neutralizing and enhancing activities of these antibodies identified an RBD-binding antibody that potently neutralized both SARS-CoV-1 and SARS-CoV-2 and did so without promoting ADE of viral entry. Interestingly, antibodies to one group of RBD epitopes were significantly associated with ADE of entry, while also exhibiting various degrees of neutralization.

Blood Components

In modern medical treatments, patients may receive a pint of whole blood or just the specific components of the blood that are needed to treat their particular condition. This approach to treatment, referred to as blood component therapy, allows several patients to benefit from one pint of donated whole blood.

The transfusable components that can be derived from donated blood are red cells, platelets, plasma, cryoprecipitated AHF (cryo), and granulocytes. An additional component, white cells, is often removed from donated blood before transfusion.

White Cells & Granulocytes

Whole blood contains red cells, white cells, and platelets (

45% of volume) suspended in blood plasma (

  • Color: Red
  • Shelf Life: 21/35 days*
  • Storage Conditions: Refrigerated
  • Key Uses: Trauma, Surgery

Whole Blood is the simplest, most common type of blood donation. It’s also the most flexible because it can be transfused in its original form, or used to help multiple people when separated into its specific components of red cells, plasma and platelets.

A whole blood donation requires minimal processing before it is ready to be transfused into a patient. If not needed right away, whole blood can be refrigerated for up to 35 days, depending on the type of anticoagulant used.

Whole blood is used to treat patients who need all the components of blood, such as those who have sustained significant blood loss due to trauma or surgery.

Whole blood can be donated at any Red Cross blood drive or blood center.

* Shelf life of whole blood varies based on the type anticoagulant used.

Red blood cells (RBCs), or erythrocytes, give blood its distinctive color. Produced in our bone marrow, they carry oxygen from our lungs to the rest of our bodies and take carbon dioxide back to our lungs to be exhaled. There are about one billion red blood cells in two to three drops of blood.

  • Color: Red
  • Shelf Life: Up to 42 days*
  • Storage Conditions: Refrigerated
  • Key Uses: Trauma, Surgery, Anemia, Any blood loss, Blood disorders, such as sickle cell

Red blood cells are prepared from whole blood by removing the plasma (the liquid portion of the blood). They have a shelf life of up to 42 days, depending on the type of anticoagulant used. They can also be treated and frozen for 10 years or more.

RBCs are used to treat anemia without substantially increasing the patient’s blood volume. Patients who benefit most from transfusion of red blood cells include those with chronic anemia resulting from kidney failure or gastrointestinal bleeding, and those with acute blood loss resulting from trauma. They can also be used to treat blood disorders such as sickle cell disease.

Prestorage Leukocyte-Reduced Red Blood Cells

Leukocyte-reduced RBCs are prepared by removing leukocytes (white blood cells) by filtration shortly after donation. This is done before the RBCs are stored because over time the leukocytes can fragment, deteriorate, and release cytokines, which can trigger negative reactions in the patient who receives them. These reactions can occur during the initial transfusion or during any future transfusions.

Donating Red Blood Cells

The Red Cross calls RBC donations “Power Red.” By donating Power Red, you double your impact by contributing two units of red cells in just one donation.

* Shelf life of red cells varies based on the type anticoagulant used.

Platelets, or thrombocytes, are small, colorless cell fragments in our blood whose main function is to stick to the lining of blood vessels and help stop or prevent bleeding. Platelets are made in our bone marrow.

  • Color: Colorless
  • Shelf Life: 5 days
  • Storage Conditions: Room temperature with constant agitation to prevent clumping
  • Key Uses: Cancer treatments, Organ transplants, Surgery

Platelets can be prepared by using a centrifuge to separate the platelet-rich plasma from donated whole blood. Platelets from several different donors are then combined to make one tranfusable unit. Alternately, platelets can be obtained using an apheresis machine which draws blood from the donor’s arm, separates the blood into its components, retains some of the platelets, and returns the remainder of the blood to the donor. Using this process, one donor can contribute about four to six times as many platelets as a unit of platelets obtained from a whole blood donation.

Platelets are stored at room temperature for up to 5 days. They must receive constant gentle agitation to prevent them from clumping.

Platelets are most often used during cancer treatment as well as surgical procedures such as organ transplant, in order to treat a condition called thrombocytopenia, in which there is a shortage of platelets. They are also used to treat platelet function abnormalities.

Since platelets must be used within 5 days of donation, there is a constant need for platelet donors.

Plasma is the liquid portion of blood our red and white blood cells and platelets are suspended in plasma as they move throughout our bodies.

  • Color: Yellowish
  • Shelf Life: 1 year
  • Storage Conditions: Frozen
  • Key Uses: Burn patients, Shock, Bleeding disorders

Blood plasma serves several important functions in our bodies, despite being about 92% water. (Plasma also contains 7% vital proteins such as albumin, gamma globulin and anti-hemophilic factor, and 1% mineral salts, sugars, fats, hormones and vitamins.) It helps us maintain a satisfactory blood pressure and volume, and supplies critical proteins for blood clotting and immunity. It also carries electrolytes such as sodium and potassium to our muscles and helps to maintain a proper pH (acid-base) balance in the body, which is critical to cell function.

Plasma is obtained by separating the liquid portion of blood from the cells. Plasma is frozen within 24 hours of being donated in order to preserve the valuable clotting factors. It is then stored for up to one year, and thawed when needed.

Plasma is commonly transfused to trauma, burn and shock patients, as well as people with severe liver disease or multiple clotting factor deficiencies.

In some cases, patients need plasma derivatives instead. These are concentrates of specific plasma proteins obtained through a process known as fractionation. The derivatives are treated with heat and/or solvent detergent to kill certain viruses like those that cause HIV, hepatitis B, and hepatitis C.

Plasma derivatives include:

  • Factor VIII Concentrate
  • Factor IX Concentrate
  • Anti-Inhibitor Coagulation Complex (AICC)
  • Albumin
  • Immune Globulins, including Rh Immune Globulin
  • Anti-Thrombin III Concentrate
  • Alpha 1-Proteinase Inhibitor Concentrate

When collecting specifically plasma, the Red Cross is seeking AB-type donors. AB plasma is collected at select Red Cross Donation Centers only.

Epitope Detection of Antigens by Antibodies | Immunology

Many clinical tests are being done on the basis of specificity of antibodies for antigen and their ability to recognize epitopes, which is a very small portion or portions of antigen. Antibody based assays are epitope-detecting devices and most of them are based upon the quantitative pre­cipitin curve (Fig. 5.7).

A.1 Precipitation reactions in fluids:

A quantitative precipitation reaction can be performed when a series of test tubes are taken with a constant amount of antibodies with which the amount of antigens are gradually increased, it starts to generate precipitate.

After the precipitate forms, each tube is centrifuged, pellet is collected by pouring the supernatant and is measured. A precipitin curve is generated by plotting the amount of precipitate against increasing antigen concentration (Fig. 5.7).

The curve shows different zones called:

ii. Zone of excess antibody

iii. Zone of excess antigen

Equivalence zone is a zone of forming large multi-mole lattice where the amount of antibody and antigen is optimal. The complex increases in size and precipitates out of solution.

The zone of excess antibody shows the pre­sence of unreacted antibody in the supernatant with some soluble complexes consists of multiple molecules of antibody bound to a single molecule of antigen.

The zone of excess antigen is that region where the unreacted antigen can be detected and small complexes are again observed where one or two molecules of antigen bound to a single molecule of antibody.

A.2 Precipitation in particulate antigens:

Particulate antigens like erythrocytes, bacte­ria or antigen-coated latex beads are normally evenly dispersed in suspension. The homo-genicity of the suspension is disrupted by the cross- linking of antigen-bearing particles by antibodies, leading towards clumping of the particles, known as agglutination.

The antibodies which produce agglutination reactions are called agglutinins, which is similar in principle with precipitation reaction. The excess amount of antibody inhibits agglutination reactions, called pro-zone effect.

When the antigen is an erythrocyte called hemagglutination or IgM antibodies efficiently cross-link the particles called direct agglutination, or whether an anti-immunoglobulin called indirect or passive agglutination or with bacterial antigen called bacterial agglutination.

This reaction normally involves IgM anti­bodies that cross link epitopes on cells or particles. IgM is the largest immunoglobulin has ten epitope- binding sites so they are less efficient in direct agglutination.

It is a regular phenomenon of ABO-blood group antigen when RBCs are mixed on a slide with antisera to the A and B blood-group antigens, may form a clump depending upon this reaction blood transfusion principle is determined (Table 5.4 and Fig. 5.8).

Bacterial agglutination:

When bacterial infection, takes place against the surface antigen present on the bacterial wall, generates serum antibodies. This reaction is obser­ved by the bacterial agglutination reaction. The agglutinin titer of an antiserum can be used to diag­nose a bacterial infection, when a patient suffers from typhoid fever, shows a significant rise in the agglutination titer to Salmonella typhi. Agglutination reactions also provide a way to type-bacteria.

Passive or Indirect agglutination:

This technique is often used to detect non-IgM antibodies or antibodies in concentrations too low to be detected by direct agglutination. Human antibodies may not directly agglutinate antigen- bearing particles. The sensitivity and simplicity of agglutination reactions can be extended to soluble antigens by the technique called reverse passive hemagglutination test.

In case of this technique RBC is treated with tannic acid or chromium chlo­ride. This helps in detection of antigen in the patient’s serum or in the blood donated by a professional donor. It is used to detect Hepatitis-B antigen in blood to be used for transfusion obtained from a blood bank. It is far more sensi­tive than precipitation reaction. It is also can be performed with antigen-coated particles of latex (Fig. 5.8).

This is also called Anti-globulin (Fig. 5.9) test. It may be direct or indirect test.

Sometimes antibodies bind to erythrocytes, do not agglutinate as because the ratio of Ag/Ab shows either excess antigen or excess antibody or electrical charges on the red blood cells, hinder the effective cross linking of the cells.

These anti­bodies are called incomplete antibody. To detect the presence of non-agglutinating antibodies on RBC, a secondary Ab can be used. After that they can cross-link with erythrocytes result in agglu­tination.

(ii) Indirect Coomb’s test:

Sometimes for particular laboratory purpose, it is necessary to know whether a serum sample has particular antibody for specific RBC. By this detection process, the presence or absence of potential non-agglutinating antibodies in the sam­ple can be detected.

In indirect coomb’s method, the RBCs with the serum sample is incubated. After that a wash is taken for removal of unbound antibodies and then a second anti-immunoglobulin component is added and it shows cross-linking with the cells.

Agglutination inhibition:

A modification of the agglutination reaction is called agglutination inhibition which is a very sensitive assessment technique which can even quantify a little amount antigen. In case of home pregnancy test kit, it includes latex particles coat­ed with human chorionic gonadotropin (HCG) and antibody to HCG.

The addition of urine sam­ple from a pregnant woman, which contains HCG, inhibits agglutination of the latex particles when the anti-HCG antibody is added, thus the absence of agglutination indicates pregnancy. Agglutination inhibition assays are widely used in clinical laboratories to determine if an individual has been exposed to certain types of viruses that cause agglutination of red blood cells.

The epitopes present on soluble molecules will precipitate upon reaction with the “right” amount of antibody. Precipitates can also form in an agar matrix. The quantitative precipitin reac­tion requires the preparation.

Single Radial Immuno-diffusion (SRID):

This technique is also called the Mancini technique after the name of the person. It is based upon the diffusion of soluble antigen within an agar-gel which contains a uniform concentration of antibody. Antibody containing molten-agar is poured into a glass slide or plastic dish, gradually cools and solidifies after that the wells are cut into the gel-matrix.

Soluble antigen is placed into the well (Fig. 5.10 and Fig. 5.11). Antigen diffuses radially from the well, forming a precipitin ring at equivalence. The area of the precipitin ring is pro-portional to the concentration of antigen. The concentration of antigen in a test sample can be accurately determined by comparing its diameter with a standard calibration curve.

The Mancini method is regularly used to quantitate serum levels of IgM, IgG and IgA by incorporating class-specific anti-iso-type antibody into the agar. The limitation of this technique is that it can not detect antigens when the concen­tration goes below 5-10 μg/ml.

Double Immuno Diffusion (DID):

This technique, also called Ouchterlony tech­nique, is based upon the diffusion of both antigen (located in one well) and antibody (located in another well) through an agar gel forming a con­centration gradient. As equivalence is reached a visible line of precipitation is formed. It is an effective qualitative tool for determining the relationship between antigen and the number of different Ag-Ab systems present.

An advantage of this technique is that several antigens or antibod­ies can be Compared to determine identity (when two antigens share identical epitopes), nonidentity (when two antigens are unrelated epitopes), partial identity (when two antigens share some epitopes but one or the other has a unique epitope or epitopes) etc. (Fig. 5.12 and 5.13).

Immuno-electrophoresis (IEP):

This technique is a modification of double diffusion which is a combination of separation by electrophoresis with identification by double immunodiffusion. Antigens are loaded into a well within the agar, an electrical current is applied and antigens migrate according to their size and elec­trical charge. The formation of precipitin bands with polyvalent or specific antiserum identifies individual antigen components (Fig. 5.14).

Immunoelectrophoresis is greatly used in clinical laboratories to detect the presence or absence of proteins in the serum and the detection of antibody concentrations qualitatively.

Future directions

Significant progress has been made in both the understanding of TRALI and its mitigation. Future strategies for the prevention of TRALI may include immune-based strategies to block donor antibodies or the immune response to transfused antibodies. Human investigations have found increased plasma levels of the inflammatory cytokine IL-8 but lower levels of the anti-inflammatory cytokine IL-10 in both antibody- and nonantibody-mediated cases of TRALI compared with those of possible TRALI. 1,20,126 Recently, it was shown that regulatory T cells and dendritic cells were protective in a murine model of antibody-mediated TRALI, and this protection was associated with increased plasma levels of IL-10. 127 In addition, there was evidence of a benefit of IL-10 administration in the prevention of murine TRALI as well as treatment after its induction with HLA antibodies (Figure 2). These findings may indicate a distinct mechanism of pathogenesis in TRALI compared with other forms of lung injury as well as opportunities for prevention and treatment.

TRALI suppression in CD4+ T cell–depleted mice with murine IL-10 administration. (A) Decreased lung wet/dry (W/D) weight ratios of CD4+ T cell–depleted C57BL/6 mice infused with 34-1-2s/AF6- and treated prophylactically with murine IL-10 administration (45 mg/kg intravenously). (B) Lung W/D weight ratios of CD4+ T cell–depleted C57BL/6 mice infused with 34-1-2s/AF6- and treated therapeutically 15 minutes later with or without murine IL-10 administration (45 mg/kg intravenously) after onset of TRALI (at least a 2° drop in rectal temperature 10 minutes after TRALI antibody injection). Comparisons in both panels were analyzed with one-tailed unpaired t test. Each dot represents 1 mouse, and error bars represent standard deviation. *P < .05 ****P <. 0001. PBS, phosphate-buffered saline. Reprinted from Kapur et al 127 with permission.

TRALI suppression in CD4+ T cell–depleted mice with murine IL-10 administration. (A) Decreased lung wet/dry (W/D) weight ratios of CD4+ T cell–depleted C57BL/6 mice infused with 34-1-2s/AF6- and treated prophylactically with murine IL-10 administration (45 mg/kg intravenously). (B) Lung W/D weight ratios of CD4+ T cell–depleted C57BL/6 mice infused with 34-1-2s/AF6- and treated therapeutically 15 minutes later with or without murine IL-10 administration (45 mg/kg intravenously) after onset of TRALI (at least a 2° drop in rectal temperature 10 minutes after TRALI antibody injection). Comparisons in both panels were analyzed with one-tailed unpaired t test. Each dot represents 1 mouse, and error bars represent standard deviation. *P < .05 ****P <. 0001. PBS, phosphate-buffered saline. Reprinted from Kapur et al 127 with permission.

Prior studies have suggested a lack of benefit of the anti-inflammatory effects of systemic corticosteroids in ARDS and a lipopolysaccharide-stimulated murine model of TRALI. 60,128 Differences in murine TRALI models or the timing of corticosteroid administration may explain these disparate effects. Other anti-inflammatory modalities to prevent or treat TRALI have also been proposed, including the targeting of C-reactive protein, IL-8, reactive oxidative species, neutrophil extracellular traps, or Fc receptors. 129 Additional investigations are needed to confirm that these murine model findings parallel human pathophysiology before embarking on clinical studies of TRALI prevention or treatment. The recognition that any immunomodulatory therapy could increase the risk of infection is especially relevant in immunocompromised patients (eg, those with hematologic malignancies). In contrast to TRALI, few in vitro or in vivo investigations have been conducted to further understand the pathogenesis of TACO. In vivo models of TACO are needed to study the effect of individual blood components relative to other intravenous fluids on pulmonary capillary pressures, specific rates of blood administration, and the benefit of prophylactic diuretics as well as the role of systemic inflammation.

Work to improve blood collection and storage may also provide the opportunity to prevent TACO and TRALI. Modifications to blood components include the development of novel filters, apheresis collection systems, pathogen reduction, extended storage of platelets, and new methods of leukodepletion or irradiation. A novel prestorage filter absorbed HLA antibodies and lipids in addition to leukocytes and platelets, and it was associated with reduced neutrophil activation and TRALI incidence in an animal model. 130 Translation of an effective filter to clinical practice could obviate the need for specific donor mitigation strategies. Continued research to develop RBC and platelet storage solutions that preserve blood product quality but also mitigate TRALI also hold promise. 131-134 In addition, an ongoing randomized clinical trial is revisiting the potential benefit of washing allogeneic RBCs in reducing the incidence of TACO and TRALI. 92,135

At a systems level, prevention of TACO and TRALI requires implementation science research to harness the ever-expanding role of EHRs in medical care. The application of natural language processing or machine learning methods to EHR data of transfusion recipients would allow for more widespread surveillance and sophisticated approaches to adverse event reporting for blood and blood-derived products. Automated identification of adverse events utilizing large sources of blood donor, component, and transfusion recipient data would allow assessment of the safety of blood component modifications (eg, pathogen-reduced products or extended storage of platelets) in addition to additional mitigation strategies. Expanded active surveillance of pulmonary transfusion reactions would also serve to aid in the refinement of current diagnostic criteria for TACO and TRALI, allowing better discrimination of individual reactions from other causes of pulmonary edema.

Predictive algorithms could also be embedded within the EHR to allow for real-time identification of patients at increased risk of an adverse pulmonary transfusion event. For example, clinical decision support systems incorporating hemodynamic parameters or creatinine clearance could trigger recommendations for diuretic administration or alternatives to transfusion in patients at risk for TACO. Evidence of a systemic inflammatory response or other recipient risk factors could trigger allocation of lower-risk platelet or plasma units to hospitalized patients at increased risk for TRALI. Real-time identification of at-risk individuals could also be coupled to clinical trials of immune-based therapies for TRALI. Continued collaboration by bench and clinical researchers, clinicians, epidemiologists, and blood donor centers will be required to further understand and minimize these severe complications of blood transfusion.


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