Modification of double color indirect immunofluorescent based platelet crossmatch for patients with anti-platelet antibodies

Abstract

© 2020, Chiang Mai University. All rights reserved. Platelet crossmatch is necessary to select the best platelets for patients with platelet antibodies. Two platforms available in special service laboratory. They are monoclonal immobilization of platelet antigen (MAIPA) and solid phase red cell adherence assay (SPRCA). However, MAIPA is relatively time-consuming and not easily adapted for routine use while SPRCA requires a high level of skill to perform the test. In contrast, flow cytometric analysis routinely shows greater sensitivity and can be developed for platelet crossmatch. This work aimed to modify double color immunofluorescence for platelet crossmatch. Two antibodies, PE-goat F (ab')2 anti-human IgG, Fc specific and FITC-mouse anti-human CD41 antibody were optimized. Treatment of platelets and sequence of the process were studied. Optimal concentrations of PE-goat F(ab′)2 anti-human IgG, Fc specific is 1 µg/ml and FITC-mouse anti-human CD41 antibody is 2 µl/reaction. Donor platelets should be fixed and blocked with 1% paraformaldehyde and heat-inactivated normal AB serum prior to any assay. Serum should be first added followed by FITC-mouse anti-human CD41 antibody and PE-goat F(ab′)2 anti-human IgG, Fc specific. Various backgrounds were observed for different ABO individual donor autologous controls. Therefore, subtracting signal between donor autologous and crossmatch tests should be performed for interpretation. The method is effectively used for platelet crossmatch without antibody screening or donor HLA typing. Donor autologous control is suggested to perform in parallel for the best interpretation.