HLA‐DRw52‐associated DRB1 alleles: Identification using polymerase chain reaction‐amplified DNA, sequence‐specific oligonucleotide probes, and a chemiluminescent detection system

Abstract

Polymerase chain reaction (PCR) primers were designed to specifically amplify exon 2 of the DRw52‐associated DRB1 alleles for subsequent typing by sequence‐specific oligonucleotide probe (SSOP) hybridization and chemiluminescent detection. The DRw52 DRB1 group, encoding 22 of the 44 W.H.O. designated DRB1 allelic products, was divided by differential PCR with two polymorphism‐directed forward primers. Based on a polymorphism at codon 13, these forward primers separate the DRw52‐associated alleles into subsets; one comprised of the alleles of DR3/DRw11/DRw6 and the other of DRw8/DRw12/DRB1*1404. The DRB1 alleles in the latter subset were then defined by SSOP hybridization to the amplified DNA. The preferential amplification also resulted in SSOP definition of 15 alleles in the DR3/DRw11/DRw6 subset but some DRw11/DRw13 heterozygous allelic combinations were still unresolved. Two reverse PCR primers specific for the polymorphism at codon 86 were used to obtain amplified material to which SSOP reactivity provided definitive identification of the ambiguous heterozygotes.