Detection and quantitation of ABO RBC chimerism by a modified coil planet centrifuge method

Abstract

A modified coil planet centrifuge method is established by which ABO chimerism could quantitatively be analyzed and ABO variants identified to the same degree of accuracy as the other differential methods and flow cytometry. The monitoring of ABO chimerism might also help the diagnosis of early relapse or rejection after ABO incompatible HPCT.

An automatable format for accurate immunohematology testing by flow cytometry

“…FC also detected 99.5 percent of clinically relevant alloantibodies, while CAT and LISS‐IAT identified 98.9 percent and 94.7 percent, respectively. To overcome the necessity of dispersing RBC agglutinates before FC, we optimized a vacuum filtration method that eliminated the need for vigorous vortexing and pipetting or caustic agents like glutaraldehyde 4,16,17,22 . Using the filter plate FC technique, 104 of 109 samples (95.4%) were correctly typed for ABO and D, whereas the remaining five samples were read as NTD.…”

“…Importantly, these comparisons were made using authentic clinical samples with abnormalities including weak alloantibodies, rouleaux, autoantibodies, and mixed‐field reactions, which are not uncommon in hospitalized patients. In contrast, other recently described methods have only used samples from healthy donors, and none of these methods has been applied to routine alloantibody identification 19–22 . We have not yet examined assay performance for pretransfusion cross‐matches, but we anticipate that accuracy will be similar to that seen for ABO serum testing (“immediate spin” cross‐match) and RBC alloantibody testing (“Coombs” cross‐match).…”

Blood doping in athletes—Detection of allogeneic blood transfusions by flow cytofluorometry

“…Use nonagglutinating (IgG) antibodies Chemically convert agglutinating IgM pentamers into nonagglutinating IgM monomers, e.g., using dithiothreitol [48,57] or 2-mercaptoethanol Purify IgG fractions of sera on Protein G [49] Dilute antisera past point of agglutination Avoid use of potentiators Chemically fix the RBCs prior to incubation with anti-A, -B, -M, or -N, e.g., with dimethylsuberimidate [74], formaldehyde [47,75], or glutaraldehyde [13,21,44,49] To avoid agglutination caused by secondary antibody:…”

“…To detect allogeneic transfusions, currently available methods are based on the antigenic differences between the recipient's RBCs and the donor's RBCs. Originally, serological techniques were performed, such as detection of ''mixed field'' reactions when performing antigen typing and differential agglutination where percentages of agglutinated and unagglutinated RBCs were determined manually or by automated methods [9][10][11][12][13]. More sensitive quantitative techniques to characterize transfused RBCs are the enzyme-linked antiglobulin test [14] and flow cytofluorometry [15][16][17][18][19][20][21][22][23].…”

“…Formaldehyde [47,75] is a satisfactory fixative for the A, B, M, and N antigens though rather tedious (a two-day, three-step process) and causes some hemolysis. Fixation with glutaraldehyde [13,21,44,49] is rapid and nonhemolytic, and although the A, B, and M blood groups are not destroyed, the N antigen is sensitive (A. Guest, Personal communication, IBGRL, Bristol).…”