Low resolution DNA typing of the HLA-B5 cross-reactive group by nested PCR-SSP

Abstract

We have established a DNA typing system for the HLA-B5 serologically cross-reactive group (CREG) by means of a two-step PCR amplification with nested sequence-specific primers (nPCR-SSP). The present study provides a low resolution definition of the HLA-B5 CREG, i.e. identifying polymorphism equivalent to serology. Two different primer combinations allow group-specific amplification of all HLA-B5 CREG alleles and other related HLA class I alleles from genomic DNA. The amplified DNA is subjected to a second amplification step using eleven nested primer pairs. This assay permits the detection of the HLA-B5 CREG specificities B35, B51, B52, B53, and B7801 in all homozygous and heterozygous combinations. Sensitivity and specificity as judged by a blind quality control study investigating a reference panel (n = 50) is 100%. Extension of this approach should allow rapid DNA typing of all serologically defined HLA-B specificities by nPCR-SSP.