Principles and Applications of Red Blood Cell Flow Cytometry

Abstract

Objective: In this review we address principles and important applications of flow cytometry in immunohematology. Sources and Criteria: Some studies of the theoretical back-ground and recent publications of current applications were selected. Results: The direct, indirect and two-color fluorescence as well as the ‘one-antigen-one-antibody’ model are addressed. Possible standardizations and quality controls are discussed. Flow cytometric analyses with erythrocytes bear special problems such as agglutination, which is of lesser im-portance with other cells types. Possible applications include the quantitation of antigen density, antibody concentration, red blood cell bound immunoglobulin, and red blood cell sub-populations. Conclusion: Red blood cell flow cytometry is very sensitive, has a high potential for automatization, and hence may gain importance especially for many routine tasks in the future.

Quantification of glycophorin A and glycophorin B on normal human RBCs by flow cytometry

“…The main problem in the flow cytometry analysis was the agglutination of RBC by the MoAbs. If agglutination occurs during the sample preparation and before the measurements, it may compromise the results because it might result in selective enrichment or loss of antigen‐positive cells 27 . All the MoAbs used caused agglutination, principally at the highest concentrations utilized.…”

Rh flow cytometry: An updated methodology for D antigen density applied to weak D types 164 and 165

“…The mixed-field reactivity, seemingly confirmed by agglutination in gel matrix tests (Figure S1), had not been reported in any of the previous observations of weak D type 18. [32][33][34] The red cells belonged to a pregnant woman and factors other than the RHD variant could have caused the mixed-field reactivity, such as hematologic pathologies and treatment 40 or pregnancy. 84 However, the woman was reported healthy without transfusion.…”

“…30 Some weak D types, such as type 10 (W393R), 31 type 18 (R7W), [32][33][34] and type 31 (P6L), 35,36 are known to present with irregular quantitative expression of the D antigen, such as mixed-field agglutination during serologic testing or as multiple red cell populations on flow-cytometric histograms. This phenomenon of mixed-field reactivity in blood group antigens has been explained over the years by several mechanisms including transcriptional noise, 37 red cell chimerism, 38,39 maternal-fetal hemorrhage, 40 or sample preparation error during flow cytometry. 41 As novel blood group alleles continue to be identified, comprehensive genetic and serologic descriptions are standard practice.…”