GENOTYPING FROM HPA-1 TO HPA-5 WITH ALLEL E-SPECIFIC LIGASE CHAIN-REACTIONS FOLLOWING AMPLIFICATION OF GENOMIC DNA IN A MULTIPLEX PCR

Platelet-specific antibodies are involved in refractoriness to platele t transfusions, neonatal alloimmune thrombocytopenia, and posttransfus ion purpura. Binding to different epitopes platelet glycoproteins, hum an platelet antigens (HPA), they can cause severe The polymorphic DNA regions of the best known HPA systems HPA-1 to HPA-5 were amplified fr om genomic DNA in a multiplex polymerase chain reaction (PCR). An auto matable ligation-based assay (LCR) using allele-specific oligonucleoti de probes was developed for rapid genotyping. The results were correla ted with the well established allele-specific restriction enzyme assay (ASRA), and the phenotype was determined by the monoclonal antibody-s pecific immobilization of platelet antigens (MAIPA). We found a 100 % concordance of genotypes between the HPA-LCR in 54 cases of NAIT, PTP and normal controls with the established methods. The gene frequencies calculated from 216 German and 55 Japanese random blood donors were s imilar to those determined by other time-consuming typing methods in a Caucasian and Japanese population, respectively. Multiplex PCR and th e allele-specific HPA ligation assay provide a reliable tool for typin g both small series and all platelet donors of a transfusion center in order to identify rare genotypes necessary for the treatment of patie nts suffering from NAIT, PTP or refractoriness to platelet transfusion s.