Sandra J. Nance

Predicting the clinical significance of red cell alloantibodies using a monocyte monolayer assay.

Few data have been published that correlate in vitro monocyte monolayer assays (MMA) and red cell (RBC) survival in patients with alloantibodies of unknown significance. Over the past 6 years we gathered clinical correlations in 12 patients with the following antibodies: anti‐Lan (three patients), ‐Ge (three patients), ‐Yta (five patients), and ‐Ytb (one patient). RBC survival was estimated using 51Cr studies in seven patients and follow‐up of transfusion of incompatible blood in the other five. Six patients with no evidence of RBC destruction had negative MMA findings (anti‐Lan [one patient], ‐Ge [two patients], and ‐Yta [three patients]). Five patients with evidence of in vivo RBC destruction had significant MMA results. The two clinically significant anti‐Lans required fresh serum to give a meaningful MMA result. One patient (anti‐Ytb) had an MMA result of borderline significance‐normal 51Cr RBC survival at 1 hour‐but a reduced T50Cr. The MMA we used appeared to predict the clinical outcome of transfusion in every patient with antibodies to high‐frequency antigens whom we tested.

Application of flow cytometry to immunohematology

A method is described for applying flow cytometry to the analysis of populations of red blood cells that have been sensitized, in vivo or in vitro, with IgG antibody. Specific instrument settings and details of the method are given with sufficient explanation for practical use in immunohematologic investigations.

Determination of zygosity using flow cytometric analysis of red cell antigen strength

A flow cytometry method, developed in our laboratory to measure red cell (RBC)‐bound IgG, was compared to the manual titration technique in the measurement of RBC antigen strength to determine zygosity. Parallel studies using antibodies to antigens in the Rh, Kell, Kidd, Ss, and Duffy systems were performed. The antisera (n = 20) were tested against five examples each of RBCs from apparent homozygotes and heterozygotes. The flow cytometry method was clearly superior, showing distinct differences, with no overlap of the ranges of results, between the reactions of RBCs from homozygotes and heterozygotes with 10 of 20 (50%) antisera. By the manual titration technique none of these sera clearly demonstrated dosage and 15 showed overlap of the ranges. It was obvious from the results that the commonly used manual titration technique for comparing the test RBCs with a single example of RBCs from a homozygote and heterozygote yielded inaccurate results.

Erythromycin-Induced Immune Hemolytic Anemia

Abstract. A 3‐year‐old female receiving Pediazole (erythromycin ethylsuccinate and sulfisoxazole) for tonsillitis and otitis media developed severe hemolytic anemia. No serum drug‐dependent antibodies could be demonstrated with an in vitro ‘immune‐complex’ method using Pediazole, pure erythromycin ethylsuccinate or pure sulfisoxazole. However, a method using red cells coated with erythromycin base showed in vitro lysis of the erythromycin‐coated red cells. This is only the second case of immune hemolytic anemia associated with erythromycin and the first where in vitro drug‐dependent hemolysis was demonstrable.

MAM: a new high-incidence antigen found on multiple cell lines

BACKGROUND: Three women have been identified with an antibody to a “new” high‐incidence antigen found on multiple cell lines.

International rare donor panels: a review

International rare blood donor panels or registries are important in the consistent availability of rare blood for patients who need this scarce resource. In countries where it has been possible to commit resources to this effort and often where the need is great, donors have been entered into a registry. The ISBT leadership recognized the importance of this very challenging inventory management activity and created a Working Party to support it. Individual countries support the WHO International Rare Donor Panel by submitting their donors’ phenotype or genotype information to be catalogued into the database. It is extremely important that this database be cultivated and grown. The contributing countries keep their list updated and supply the blood product as they can when requested. It is known that some blood types are extremely scarce worldwide and requests for these are particularly difficult to fulfil. Thus, it is important to have a protocol to identify and recruit donors with rare blood types. It is equally or perhaps more important to ensure that the patients who need the rare blood are being managed appropriately in the presence and absence of rare blood products being available.

Global definitions of rare donors

A basic definition of a rare donor unit is that unit not available for the patient at the time of the transfusion need. However, there are some patients who required exquisitely phenotyped, even genotyped units, for optimal survival of transfused red cells. Often, the rare phenotyped unit needed is negative for an antigen of high prevalence, sometimes the type is less than one in 10 000 donors tested or even rarer. Further complications arise with consideration of ABO/Rh type or the concomitant presence of antibodies to common antigens. Another type of rare unit is one that is negative for multiple common antigens, which in combination make the donor difficult to find.

Flow cytometry related to red cells

Researchers in Transfusion Medicine Science have benefited from the use of the flow cytofluorometer. The flow cytometer has distinct advantages over visual examination of antigen-antibody reactions. The flow cytometer measures fluorescence per cell, and through the use of anti-IgG tagged with a fluorochrome, cells with differing levels of cell-bound IgG can be quantitated. This has been used in the study of allo- and autoantibodies. Immunohematologists, with the wide range of red cell alloantibodies in many blood group systems, have a seemingly unending supply of materials to enable studies of red cells. This article describes the published reports involving flow cytometry related to red cells. Four areas are discussed: detection of red cell-bound IgG, detection of red cell immunogobulins other than IgG, detection and quantitation of red cell antigens, and detection and quantitation of red cell populations.

Recurrent Donath-Landsteiner hemolytic anemia: a pediatric case report

Paroxysmal cold hemoglobinuria (PCH) is a form of autoimmune hemolytic anemia caused by the Donath‐Landsteiner antibody (D‐L antibody). In children, this is typically a transient immune‐mediated hemolysis that follows a viral illness and does not recur. Recurrent acute or chronic PCH due to D‐L antibody is very rare.

Preparation of reticulocyte‐rich red cells using LeucoPREP

To the Editor: Our data’ and those of Jones et al.’ clearly demonstrate that false-positive reactions do occur between chemically-modified anti-D and Rh-negative red cells that are non-reactive with anti-A and/or anti-B. In the absence of tests with an immunologically inert Rh control reagent, the results of tests with anti-D may be misinterpreted. Our concerns with the paper by Jones et al.’ related to lack of specific procedural information, their apparent use of highprotein anti-D without an immunologically inert control reagent (a practice that is ~ontraindicated),~ absence of data on the observed strength of their false-positive tests, and their conclusion: namely, that a separate immunologically inert control reagent is essential to avoid misinterpretation of results obtained with chemically-modified anti-D. In our opinion, there are better ways to ensure that proper conclusions are made. Despite the absence of emphasis on grading of serologic reactions, as exemplified by lack of specific comment in reagent package inserts and the AABB Technical M a n ~ a l , ~ we believe that serologic reactions should be graded and the results interpreted accordingly. Although weak reactivity may be a reflection of antigen expression, such results also may be due to dysproteinemia, precoating of red cells with globulins (as in autoimmune hemolytic anemia), or red cell adherence to leukocytes or fibrin. These latter reactions are usually weak but may be enhanced in reagent diluents containing substances that potentiate hemagglutination. Thus, requiring that a minimum degree of reactivity be observed before blood typing tests are interpreted as positive is a cost-effective means to minimize errors in interpretation. While grading of serologic reactions is highly subjective, we would be exceedingly uncomfortable using a reagent anti-D that yielded reactions of 2 + or less with 20% of samples. Such a product barely meets FDA requirement^.^ Reagent manufacturers’ product directions indicate that optimal use of chemicallymodified anti-D necessitates brief incubation of tests before centrifugation in order to promote clear-cut 3+ and 4+ reactions. In our experience, use of serum suspended red cells rather than washed red cells also potentiates reactivity. In our study, using serum suspended red cells, we found the incidence of weak (2+ or less) reactions to be 0.02% of all samples tested, a frequency that involves only occasional additional investigations. Comparison of existing records with current results is required by AABB standard^,^ as an ongoing quality assurance measure. We find that approximately 50% of samples are from patients without records of previous Rh typing. We argue that it costs little more to retest these with a second anti-D and Rh control, than it does to test all samples with chemically modified anti-D and a separate Rh control, as suggested by Jones et a1.* The latter necessitates performing 2 X N tests. Testing the same number (N) of samples with chemically modified anti-D (1 X N tests) and then retesting half of those (0.5N) samples with an additional anti-D and Rh control (0.5 X N X 2) also necessitates 2N tests. The only difference in cost relates to price differences between anti-D and Rh control reagents; we consider the minimal additional money well spent. Moreover, we also argue that our protocol is less costly than testing all samples with two anti-D reagents (and now presumably a separate Rh control) as apparently practiced by Jones et al.’ W. JOHN JUDD, FIMLS, MIBIOL E. ANN. STEINER, MT(ASCP)SBB HAROLD A. OBERMAN, MD Universig of Michigan, Ann Arbor, MI