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Introduction: Patients with hematological and oncological malignancies are among the most frequent recipients of chronic red blood cell (RBC) transfusion therapy. One of the risks associated with chronic RBC exposure is the development of blood group antibodies. However, there are few extensive studies of alloimmunization rates associated with hematological or oncological malignancies in adult patients despite the frequency with which they are transfused. As such, the aims of this study were: 1) to determine the alloimmunization rate associated with malignant conditions in general, 2) to establish and compare alloimmunization rates for individuals with different forms of hematological and oncological neoplasia, and 3) to determine if any oncological disorder(s) appeared to predispose individuals to alloimmunization such that antibody mitigation measures could be employed for these diagnoses. Methods: Patients included were those undergoing type and screen testing at the study site (n=18,750 unique, active subjects in our facility's transfusion record database). For each of these subjects, the electronic medical record was retrospectively reviewed to determine if a history of a hematologic or oncologic malignancy was present. Disorders grouped as 'hematologic malignancies' in this study included: acute leukemia (myeloid or lymphoid), mast cell diseases, multiple myeloma, myelodysplastic syndromes, myeloproliferative disorders, and non-Hodgkin lymphoma (T- or B-cell). 'Non-hematologic' cancers in this study were broadly grouped as follows: bladder, colon, head/neck, lung, prostate, skin, soft tissue/sarcoma. If a patient had a history of any of the above neoplastic diseases, further inclusion in the study required: 1) that the patient undergo at least one RBC unit transfusion after their diagnosis was established and 2) that at least one follow-up antibody screen test be performed after their first RBC transfusion post-diagnosis. When these additional inclusion criteria were met, the following data were collected for each subject: malignancy type, race/ethnicity, gender, and the number and specificity of antibodies detected. Ratio data were compared using the chi-square test with Yates' correction for continuity; P values Results: A total of 69 patients became alloimmunized after their diagnosis of a malignancy; these patients were overwhelmingly male (66/69; 95.7%) and most identified as white, non-Hispanic (50/69; 72.5%). The alloimmunization rate among patients with any malignancy diagnosis was 1.5% (69/4687), which was significantly lower than the general, historical alloimmunization rate at our facility (443/18750; 2.4%; P=0.0019). Of total transfused patients with a malignancy, most were diagnosed with a non-hematological disorder(4371/4687; 93.3%) and such malignancies were associated with an alloimmunization rate of 1.3% (55/4371) following transfusion. By comparison, the alloimmunization rate for transfused patients with hematological malignancies was 2.8% (14/501), which was significantly higher (P=0.011). When alloantibody development was analyzed on a disorder-by-disorder basis, the five highest alloimmunization rates were seen in: myelodysplastic syndromes (8/71; 11.2%), soft tissue cancers (3/50; 6.0%), acute leukemia (1/34; 2.9%), renal cancer (3/113; 2.7%), and myeloproliferative disorders (2/75; 2.7%). Alloimmunization rates under 2% were observed for all other hematological and non-hematological malignancies encountered in our patient population. Conclusions: RBC alloimmunization rates in transfused patients with malignancy were significantly lower than historical controls, suggesting that the immunosuppressive nature of these disorders and/or their associated therapies may play a role in limiting blood group antibody development. Hematologic disorders were associated with higher alloimmunization rates than non-hematologic malignancies, which may reflect a larger transfusion burden in these illnesses (a factor to be evaluated in future analyses). Overall, only myelodysplastic syndromes and soft tissue malignancies were associated with antibody formation rates substantially higher than historical controls. Therefore, consideration for prophylactic antigen matching to prevent alloimmunization may be warranted for these conditions. Disclosures No relevant conflicts of interest to declare.

The presence of residual donor red cells in transfused patients impairs the ability to obtain a reliable phenotype for subsequent allogeneic blood transfusions. Post transfusion phenotyping is particularly important in transfused patients developing multiple or weak blood group antibodies. However, separation of autologous and transfused red cells from recently transfused patients is necessary for the proper identification of any suspected red cell alloantibody or autoantibody. The standard technique for the separation of autologous from transfused red cells involves microhematocrit centrifugation with phytate ester solution, a laborious and time consuming method. We have examined the effectiveness of a simplified microhematocrit technique, omitting the use of esters, to separate autologous and transfused cells by performing Rhesus (Rh) and Kell (K) blood group phenotyping, both manually and by automation, pre and post transfusion. The Rh phenotype and K type were recorded on pre- and post-transfusion peripheral blood samples from 100 newly transfused operating theatre patients, both manually and by automation using the DiaMed Classic ID-Gel Station and Diaclon ’Rh subgroups+K’ gel cards. On post transfusion blood samples, separation of autologous from transfused cells was performed using the Haematospin 1300, based on the difference in the specific gravity between reticulocytes and mature red cells where the autologous cells concentrate in the top layer of the microcapillary tube. This separation was performed on blood samples taken three days post transfusion, based upon the normal reticulocyte maturation time of three days. On all samples tested, there was 100% correlation between manual and automated ID-GEL Station results. On Rh phenotyping of post transfusion blood samples prior to cell separation, 46 of the 100 patients had a mixed field (Mf) reaction. Repeat Rh phenotyping of day three post transfusion samples, following separation of autologous cells using the microhematocrit centrifugation technique, showed complete correlation with pre transfusion Rh phenotyping results in all but two cases, where a Mf reaction persisted. These two cases had had massive blood transfusions (40 and 36 units of red cells respectively) compared to the amount of red cells (mean 3.3 units, range 1–15) transfused to the other 98 patients. One week after initial Rh phenotyping, allowing sufficient time for autologous reticulocyte recovery in these two patients, repeat Rh phenotyping of separated autologous cells correlated with that of the pre transfusion results. 6 of the 100 patients were K positive on pre transfusion testing. Repeat testing of post transfusion blood samples prior to cell separation showed a Mf reaction for the K antigen in two of these 6 cases, suggesting transfusion of K antigen negative red cell units. Once cell separation was complete, both patients were confirmed as K positive. In conclusion, the simplified microhematocrit centrifugation technique and automated ID-Gel station are reliable, sensitive and less hazardous in providing an accurate Rh phenotype and K type on patient blood samples taken three days post transfusion and could be easily performed in the standard blood transfusion laboratory. The only limitation to this technique is an inability to separate autologous and transfused cells from patients who have undergone massive transfusions within a short space of time.

It has been shown that red cells sensitized with anti-Rh antibody are agglutinated by anti-γ-globulin sera, regardless of the "order" of Rh antibody activity. Red cells sensitized with anti-Lewis antibody, on the other hand, are agglutinated by anti-sera to non-γ-globulin substances. Of six other blood group antibodies studied, five provided γ-globulin red cell coating materials and one, an anti-Kidd, a non-γ-globulin material. The participation of complement, or complement-like, substances was shown in some instances. Various pitfalls in the technics employed in absorbing Coombs sera so as to render them immunochemically specific are discussed.