Allele‐specific restriction analysis of human platelet antigen system 4

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Abstract
Background: Diagnosis, and occasionally treatment, of disorders involving platelet-specific alloimmunization, including neonatal alloimmune thrombocytopenia and posttransfusion purpura, requires platelet allotyping. Allele-specific restriction analyses for convenient genotyping are available for the major human platelet antigen (HPA) systems except HPA-4. Study Design and Methods: An allele-specific restriction analysis for HPA-4 was developed by designing a polymerase chain reaction primer containing a single-base substitution (A for T) at glycoprotein IIIa cDNA position 529. The resulting mismatch did not interfere with DNA amplification. Digestion of the polymerase chain reaction product with BsmI allowed differentiation of HPA-4a and HPA-4b alleles. Results: A 126-bp polymerase chain reaction product was amplified with the novel primer. BsmI endonuclease cleaved product encoding HPA-4a into 104- and 22-bp fragments and left DNA encoding HPA-4b intact. Conclusion: This rapid allele-specific restriction analysis for genotyping the HPA-4 system complements similar methods for other platelet alloantigenic determinants.