Expression of RhD antigen on RBC of different RhD serotype by flow cytometry

This study was purposed to establish the method of quantifying RhD antigen on red blood cells(RBC) by flow cytometry(FCM) and to explore the expression of D antigen on RBC of different RhD serotype.RhD(+) RBCs and RhD(-) RBCs were mixed in 1∶1 ratio.Cells were stained by the indirect method(IgG anti-D as the first antibody,FITC-anti-IgG F(ab′)2 as the second antibody),and the ratio of RhD(+) on RBCs was quantified by FCM.The optimal dosage of IgG anti-D was defined.Expression of RhD antigen on RBC of RhD(+),weak D,RhDel and RhD(-) type were detected by FCM.The results showed that optimal dilution of IgG anti-D monoclonal antibody was 1∶4,1×106 cells/50 μl.The percentage of D(+) RBC of RhD(+),weak D,RhDel and RhD(-) type were 96.8±2.97%,79.5±9.88%,47.8±11.43%,3.7±2.96%,respectively.The mean fluorescence intensity(MFI)of RhD antigen expression of RhD(+),weak D,RhDel and RhD(-) type were 33.3±6.21Dal,18.6±5.39 Dal,7.10±1.17Dal,0.79±0.55Dal,respectively.In conclusion,there are significant differences of RhD antigen expressions among RBC of different RhD serotypes.The level of antigen on RhD(+) RBC is the highest and then weak D the next,while the leve of antigen on RhDel RBC is the lowest level.