Pre-transfusion testing problems caused by anti-lymphocyte globulin and their solution

Anti-lymphocyte globulin (ALG) is an antibody to human lymphocytes used to decrease T-cells in renal transplant patients. We recently encountered serologic problems in testing blood from patients treated with ALG. Thirty-nine patients undergoing acute kidney rejection developed positive direct and indirect antiglobulin tests following the administration of equine ALG. Sera from these patients reacted with all red cells (RBCs) tested using both polyspecific and monospecific anti-IgG anti-human sera. Eluates prepared from the patients' RBCs showed similar reactivity. The ALG panagglutinin did not react by manual hexadimethrine bromide (Polybrene) technique. The ALG panagglutinin could be neutralized by anti-human globulin. In our hands, these techniques were useful in distinguishing ALG panagglutinin from co-existing alloantibodies.     

Transfusion problems in renal allograft recipients. Anti-lymphocyte globulin showing Lutheran system specificity

Heterologous anti-lymphocyte globulin (ALG) is known to contain panagglutination activity and may cause a positive direct antiglobulin test in a renal allograft recipient. Three patients are reported in whom the passively acquired antibody showed apparent specificity in the Lutheran blood group system. The sera of these patients and eluates from their red cells displayed reactivity consistent with a Lutheran-related antibody. While the neat ALG acted as a panagglutinin, the Lutheran system specificity appeared only following dilution of the ALG. This specificity did not appear to represent a separate, distinguishable antibody in the ALG product. Before considering the utilization of units of the rare Lu(a-b-) phenotype for transfusing a renal allograft patient who has a Lutheran-related antibody, investigation of ALG as a source of that antibody should be conducted.     

The detection of cell-bound antibody on complement-coated human red cells

This study sought to elucidate the mechanism by which human red cells, in a variety of clinical settings, become coated in vivo with autologous complement components in the absence of anti-red cell autoantibodies demonstrable by standard methods. By means of a newly developed complement-fixing antibody consumption test, previously undetectable red cell-bound gammaG globulin could be detected and quantified. By this technique, the complement-coated red cells of 13 of 16 patients were shown to carry abnormally high numbers of gammaG molecules per cell, which were nevertheless below the level for detection by the direct antiglobulin test. Eluates were made from the red cells of seven of these patients and each eluate, when sufficiently concentrated, was capable of sensitizing normal human red cells (with gammaG antibodies) to give a positive indirect antiglobulin test with anti-gammaG serum. In the presence of fresh normal serum, six of the eluates so tested were capable of fixing complement to normal human red cells. The antibodies in the red cell eluates did not exhibit Rh specificity and did not react with nonprimate red cells. When studied by sucrose gradient ultracentrifugation, the gammaG antibodies to human red cells in these eluates sedimented in the 7S region. It is concluded that in many patients in whom direct antiglobulin tests reveal only cell-bound complement, the complement fixation is mediated in vivo by small quantities of "warm-reacting" erythrocyte autoantibodies of the gammaG class.     

Comparison of antibody specificities of four anti-thymocyte/anti-lymphocyte globulin products

The antibody specificities of four anti-thymocyte/anti-lymphocyte globulin (ATG/ALG) products were examined by using a competitive immunofluorescence assay as a first step in clarifying the mechanism of ATG/ALG action. Lymphoglobuline and thymoglobuline had similar specificities with the exception of reactivity to CD28 antigen. These two ATG products blocked anti—T-cell antibodies and anti—LFA-1α antibody, while they did not react with natural killer—cell and stem-cell antigens. Antibody specificity of ATG-Fresenius was also similar to that of lymphoglobuline and thymoglobuline, except that it did not contain anti-CD4 antibody. Ahlbulin did not inhibit the binding of anti—T-cell monoclonal antibodies. Analysis of antibody specificities contained in ATG/ALG products is necessary to define specificity or combinations of specificities that are responsible for the clinical effectiveness of ATG/ALG.     

Hemolysis following intravenous immune globulin therapy

Two patients who had hemolysis after receiving large doses of intravenous immune serum globulin are reported. Both patients had positive direct antiglobulin tests due to alloantibodies contained in the immune serum globulin. Markedly increased red cell transfusion requirements and elevated serum bilirubin levels provided evidence of hemolysis.     

The efficacy of performing red cell elution studies in the pretransfusion testing of patients with positive direct antiglobulin tests

In order to evaluate the efficacy of performing red cell elutions in pretransfusion testing, the serologic records of 638 patients with positive direct antiglobulin tests (DAT) were reviewed. These patients were identified by routine antibody screening procedures that included an autologous control. DAT results on the red cells of these patients showed 279 with IgG and C3d sensitization, 319 with IgG alone, and 40 with C3d sensitization alone. Of 638 patients' red cell eluates, 401 demonstrated no reactivity, 154 demonstrated panagglutination, and 60 demonstrated passively acquired anti-A,B. Only 23 of 638 patients had alloantibody sensitization of their red cells. Of the 23, 19 had serum antibody corresponding to the specificity of antibody detected in the eluate. Thus, only four of 638 (0.6%) eluates gave results unavailable by serum testing alone. This study indicates that routine eluate investigation provides little useful information in assuring compatibility. Serum antibody testing and careful review of the clinical and transfusion history constitute appropriate pretransfusion testing in patients with positive direct antiglobulin tests. Eluate testing should be restricted to cases in which immune hemolysis is suspected clinically.     

Clinical significance of the anti-complement component of antiglobulin antisera

The need for anti-complement (anti-C') activity in antiglobulin antisera (AHG) for the detection of clinically significant antibodies was evaluated during a three-year period. While performing routine compatibility testing using standard blood banking procedures, eight patients were found whose antibodies were detectable primarily or only by AHG containing anti-C' activity; monospecific anti-igG AHG gave weak or negative reactions. Seven of the antibodies were anti-jka or jkb. Two of the anti-jka antibodies were responsible for clinically unsuspected delayed hemolytic transfusion reactions. The anti-jkb antibody resulted in a shortened survival of incompatible 51Cr-labelled red blood cells. The incidence of such "complement-only" Kidd antibodies was 23 percent of all Kidd antibodies found. These data suggest that the omission of anti-C' in AHG in routine compatability testing could result in substantial risk of failure to detect clinically significant antibodies.     

Routine Compatibility Testing Standards of the AABB as Applied to Compatibility Tests

The results of a blood group antibody survey carried out with the cooperation of 20 blood grouping laboratories are given. These results are used in a discussion of the antiglobulin phase of compatibility tests as they are outlined in the AABB Standards for a Blood Transfusion Service. A total of 37,961 irregular antibodies were reported. 37,811 or 99.6% of these antibodies could be detected by routine screening procedures using a pool of red cells from two or three selected donors, provided the pool contained the red cell antigens present in more than 2.5% of the Caucasian population. Only 145 or 0.4% of the reported antibodies would not be detected by such a routine screening test due to the fact that they react with red cell antigens present in 0.5% or less of the Caucasian population. Based on the antibody survey and a report of the antibodies responsible for 222 hemolytic transfusion reactions reported from 18 of the laboratories, a case is made for the use of routine antibody screening before all crossmatching procedures. If a carefully controlled screening test is negative it seems unnecessary to carry out the antiglobulin phase of the crossmatch. The serum or saline and high protein phases are still obligatory. If the screening test is positive, the antibody can be identified, making possible selection of suitable donor bloods before crossmatching is undertaken.     

异基因造血干细胞移植后并发自身免疫性溶血性贫血患者产生类抗体血清学检测方法及输血策略

目的探讨异基因造血干细胞移植（AHSCT）后免疫介导的自身免疫性溶血性贫血（AIHA）患者交叉配血不合的原因,血浆中不规则抗体筛选试验阳性时,不规则抗体鉴定的血清学检测方法的选择,处理措施及输血策略。 方法一例AHSCT后的3岁男性患儿因重度贫血为改善贫血症状需输注红细胞入住四川省人民医院。通过盐水试管法确定了患儿ABO、Rh血型后进行交叉配血试验发现患儿与多个同型供者血液不相合,考虑到患儿贫血严重,于是采用直接抗球蛋白试验来判断红细胞是否被致敏;通过盐水介质试管法、微柱抗球蛋白法和聚凝胺法进行ABO血型系统以外的不规则抗体筛选试验以确定血浆中有无不规则抗体;根据不规则抗体筛选试验结果选择聚凝胺法来确定抗体的特异性;通过盐水介质试管法、经典抗球蛋白法和聚凝胺法进行交叉配血实验,结合抗体鉴定的结果,综合分析选择合适的供者红细胞输注。 结果本例患儿ABO血型为AB型,Rh分型为CCDee,ABO血型已转变为供者血型。直接抗球蛋白试验强阳性,红细胞被抗体致敏。血浆中检出了不规则抗体,红细胞上放散下来的致敏抗体与血清中检出的不规则抗体均为类抗-Ce自身抗体。选择了避开类抗-Ce抗体的Ce抗原阴性的AB型红细胞输注后血色素升高,3 d后复查血常规Hb为79 g/L,输血有效。 结论任何类型的AHSCT后都有可能发展成AIHA,也就是血浆中可能存在某种自身抗体（类抗体）而破坏自身红细胞,若能够明确患者血清中存在的类抗体,即可避开这种抗体而筛选红细胞无相应抗原的供者,也就避免发生溶血性输血反应以安全输血。     

Anti-H antibody of unusually high titer showing variable reactivities against group A red cells and broad thermal amplitude in a patient with lymphoma

We report a case of a patient with high titer anti-H antibody showing broad thermal amplitude and variable reactivities against group A red cells. A 62-year-old Korean female was diagnosed with diffuse large B cell lymphoma involving multiple organs. Her ABO/RhD type was A+ and her genotype was ABO*A.01.01/ABO*O.01.02. Antibody screening test (AST) and antibody identification test (IDT) were strongly positive for all reagent cells. Anti-human globulin (AHG) test revealed an antibody titer of 1:256 for 37?°C phase and trace positivity for poly- and mono-specific C3d. Reactivity was stronger for O+ red cells than that for A+ red cells across all temperatures tested (4?°C, room temperature (RT) and 37?°C). This was also found for AHG phase. Anti-IH was ruled out based on agglutination of O+ cord cells (CCs). Antibody was determined as IgM anti-H after DTT treatment. Three batches of 10 A+ red cells from random donors were tested with three consecutive serums for crossmatching using tube method. Interestingly, out of thirty A+ red cells tested, 20 cells at RT, 11 cells at 37?°C and 11 cells in the AHG phase showed reactivity of greater than 2+. The patient was transfused with 6 units of packed RBCs subsequently. Chemotherapy (R-CHOP regimen) and Helicobacter pylori eradication were then started. Her antibody titer gradually decreased following such treatment. In conclusion, we identified a case of patient with high titer anti-H with broad thermal amplitude, suggesting that anti-H antibodies might need to be considered for cases with pan-agglutination in AST and IDT.     

Detection and quantitation of fetal maternal hemorrhage utilizing an enzyme-linked antiglobulin test

Existing methods to evaluate fetal-maternal hemorrhage depend upon red blood cell agglutination or blood film elution techniques. These tests are insensitive and difficult to quantitate and reproduce. An enzyme-linked antiglobulin test was evaluated to determine its suitability for clinical testing of postpartum candidates for Rh immune globulin administration. Prepared mixtures of Rh positive fetal and Rh negative adult red blood cells approximating fetal maternal hemorrhage ratios of 0-2.0 percent were studied. In 43 assays, the enzyme-linked antiglobulin test consistently detected Rh positive fetal red blood cells in the 0.5 and 0.25 percent mixtures representing a 25 ml and a 12.5 ml hemorrhage, respectively, in a 70-kg woman. The 0.125 percent red blood cell suspension was positive in 85 percent of the assays and the 0.0625 percent suspension was positive in 56 percent of the tests. Agglutination testing by Du variant technique failed to detect 25 percent of the 0.5 percent mixtures. Only 45 percent of tests with the Rh immune globulin crossmatch detected the 0.5 percent mixture. A modified Kleihauer-Betke procedure was as sensitive, but less reproducible than the enzyme-linked antiglobulin test. Forty-seven Rh immune globulin candidates were studied to assess the quantity of fetal maternal hemorrhage. Fourteen patients (29.8%) had detectable Rh positive red blood cells by enzyme-linked antiglobulin tests but all hemorrhages were less than 12 ml; agglutination tests did not detect any fetal red blood cells. We conclude that the enzyme-linked antiglobulin test is a simple, sensitive, and objective procedure for detecting small amounts of Rh positive red blood cells in Rh negative blood and should be applicable to clinical testing of post-partum Rh immune globulin candidates.     

Positive direct antiglobulin test results after intravenous immune globulin administration

The authors report a case of passive red cell sensitization caused by antibodies received during an infusion of intravenous immune globulin. The patient had a positive direct antiglobulin test but showed no signs of hemolysis. The intravenous immune globulin product contained antibodies against four red cell antigens. These adverse reactions should be considered in the laboratory evaluation and clinical care of patients who receive intravenous immune globulin.     

A new solid-phase assay system using magnetic force on blood group serology

Summary. A new solid-phase assay system has been developed for the detection of irregular antibodies in plasma and serum specimens. By means of a magnetic indicator cell, we have established a sensitive and easy-to-handle antihuman globulin (AHG) test involving magnetic-mixed passive hemagglutination (M-MPHA). Comparative studies of M-MPHA with conventional AHG tests demonstrated that this is sufficiently sensitive and specific to detect irregular antibodies to red blood cells. Elimination of centrifugation in this test seems to open the way toward new standardization of solid-phase systems.     

Red cell sensitization due to anti-D in anti-lymphocyte globulin

Four Rh-positive patients with severe aplastic anemia received equine anti-lymphocyte globulin. Each developed a positive direct antiglobulin test. Anti-D was identified in eluates prepared from the patients' sensitized red cells. The administered lot of anti-lymphocyte globulin was found to contain anti-D. Red cell sensitization due to passively acquired Rh antibodies can result from the administration of anti- lymphocyte globulin.     

类抗Ce自身抗体的分析与研究一例

目的通过一例类抗Ce案例的分析研究,探讨类抗体患者血清学检测方法和输血原则。方法采用盐水介质法、抗人球法进行血清学试验包括抗体筛选、抗体鉴定、直抗试验、交叉配血试验等。结果患者血清中存在类抗Ce的自身抗体。结论自身免疫性溶血的患者产生的类抗Ce抗体,在缺乏相应抗原刺激时可能快速衰减。     

抗-CD47单克隆抗体对输血前检测试验的干扰及处理措施——附1例报告

目的探讨抗-CD47单克隆抗体对输血前检测试验的干扰及处理措施。方法收集1名接受抗-CD47单克隆抗体治疗的骨髓增生异常综合征受试者标本,进行ABO和Rh血型抗原鉴定,直接抗球蛋白试验,不规则抗体筛选和抗体鉴定试验,交叉配血试验;收集多人份O型献血者血小板制备成压积血小板,与受试者血浆进行吸收试验;收集Rh抗原分型分别为CCDee、ccDEE和ccdee的O型红细胞各1人份,分别与受试者血浆进行吸收试验;使用缺乏IgG4的抗球蛋白试剂Gamma-clone进行抗体筛选与交叉配血试验,使用Immucor Capture-R固相凝集试剂盒进行不规则抗体筛选试验。结果受试者直接抗球蛋白为阳性,游离抗体筛选和鉴定试验在所有介质中均为阳性(3+~4+);血浆与红细胞多次吸收后,抗体筛选和抗体鉴定为阴性;与多次血小板吸收后,抗体筛选和抗体鉴定仍为阳性;使用Gamma-clone抗球试剂进行不规则抗体筛选和配血试验,结果均为阴性;Immucor Capture-R固相凝集试剂盒进行不规则抗体筛选试验,结果为阴性。结论抗-CD47单克隆抗体可干扰输血前检测及交叉配血,使用缺乏检测IgG4的抗球蛋白试剂Gamma-clone和Immucor capture-R固相凝集法可较好去除抗-CD47干扰。     

The Use of Papain and Bromelin in Blood Bank Screening Procedures: An Evaluation and Comparison with Standard Technics

A comparison of 1-cysteine activated papain and bromelin with saline, albumin and anti-human globulin (AHG) technics in the detection of antibodies to red cells indicated that the enzyme technics are more sensitive than saline and albumin technics; are not significantly different from the AHG technic in the detection of anti-D, anti-C and anti-e; are more sensitive in the detection of anti-E, anti-c and immune anti-A; are less sensitive in the detection of anti-Kell, anti-Duffy, anti-M and anti-S—the corresponding antigens of which they alter to prevent a positive AHG reaction. Serum inhibition of papain was prevented by the use of 1% papain up to a dilution of 1:32; enhancement of titers resulted from the use of 0.1% enzyme thereafter. A 15-minute incubation period was found to be optimal. Bromelin solutions were found to be much more stable than papain. It was concluded that although these enzyme technics are simple, economical and satisfactory within the limits applying to any enzyme technic, they are valuable adjuncts only and cannot replace the AHG test in prenatal and pretransfusion antibody screening.     

Anti-A and/or anti-B is not detectable in some patients who underwent ABO-incompatible bone marrow transplantation

BACKGROUND: Recently, anti-A and/or anti-B produced by B cells from donor marrow could not be detected for more than 20 weeks in some patients who had undergone ABO-incompatible bone marrow transplantation (BMT). STUDY DESIGN AND METHODS: Twelve to 72 weeks after 11 patients underwent ABO-incompatible BMT, titers of anti-A and anti-B were assayed, A and B antigens were identified by routine methods and flow cytometry, direct and indirect antiglobulin tests were performed, and the red cell antibody was eluted. RESULTS: In some patients who underwent ABO-incompatible BMT, anti-A and/or anti-B produced by the B cells from the donor marrow could not be detected after BMT when red cells taken from the patients before BMT carried the corresponding antigen–that is, when hematopoiesis had already changed the cells to the donor's type according to ABO blood typing. Furthermore, some blood samples from those patients gave positive results in direct antiglobulin tests. Blood typing of patients after BMT showed mixed- field agglutination. In one patient, the half-life of red cells assayed with 51Cr was 22.4 days (30.0 +/− 4.0 days for normal controls). CONCLUSION: Although many hypotheses could be considered to explain the present data, the possibility is proposed that anti-A and/or anti-B in the sera must have been consumed in some patients who underwent ABO- incompatible BMT. This may lead to problems such as difficulty of ABO typing, positive direct antiglobulin tests, and a relatively short life span of red cells.     

Role of the crossmatch in testing for serologic incompatibility

Nine unexpected antibodies of unquestioned clinical significance were detected when the major crossmatch was performed on 31,320 pretransfusion blood samples from 8969 patients whose screening test for unexpected antibodies was nonreactive. Three of the antibodies retrospectively were found to manifest a positive screening test. Another antibody was not detected by the antibody screening test due to an error in preparation of the screening red blood cells. The overriding importance of the major crossmatch is the assurance of ABO compatibility between donor blood and recipient. Therefore, while this study does not resolve whether the antiglobulin phase of the procedure might be considered optional, the major crossmatch should not be eliminated.     

肾移植后不同抗排斥治疗方案感染及急性排斥发生率的比较

背景:目前对肾移植围手术期采用何种免疫抑制治疗方案(特别是抗体的使用)既可以减少急性排斥反应,又不增加受体感染的风险尚无统一的认识。目的:比较肾移植后6个月内6种不同免疫抑制诱导治疗方案的感染及急性排斥发生率差异。方法:采用前瞻性队列研究设计,将113例同种异体肾移植后患者分为甲基泼尼龙组、甲基泼尼龙+抗人T细胞免疫球蛋白组、甲基泼尼龙+抗胸腺细胞球蛋白组、甲基泼尼龙+赛尼哌组、甲基泼尼龙+抗胸腺细胞球蛋白+赛尼哌组、甲基泼尼龙+抗人T细胞免疫球蛋白+赛尼哌组,各组免疫抑制诱导治疗后均联合环孢素A+吗替麦考酚酯+泼尼松三联维持免疫抑制治疗。结果与结论:①感染发生率:甲基泼尼龙+抗胸腺细胞球蛋白组最低,甲基泼尼龙+抗胸腺细胞球蛋白+赛尼哌组最高。各组间差异无显著性意义。②急性排斥发生率:甲基泼尼龙组最高,甲基泼尼龙+抗胸腺细胞球蛋白+赛尼哌组最低。甲基泼尼龙组明显高于甲基泼尼龙+抗胸腺细胞球蛋白+赛尼哌组及甲基泼尼龙+抗人T细胞免疫球蛋白+赛尼哌组(P<0.05)。表明联合单克隆与多克隆抗体免疫抑制诱导治疗方案与单用甲基泼尼龙组或单独使用单克隆或多克隆抗体治疗组比较,不增加感染的发生,且急性排斥反应发生率较低。