DISCRIMINATION OF HUMAN HLA‐DRB1 ALLELES BY PCR‐SSCP (SINGLE‐STRAND CONFORMATION POLYMORPHISM) METHOD

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Abstract
A single‐strand conformation polymorphism (PCR‐SSCP) method has been adopted for discrimination of human HLA‐DRB1 alleles. This method enabled the detection of DN A polymorphisms including point mutations at a variety of positions in the DN A fragments of the HLA‐DRB1 gene. A total of 27 HLA‐DRB1 alleles from 172 healthy donors were analysed using a combination of PCR‐SSCP with group‐specific amplifications. Application of a small amount of amplified and denatured DNA to non‐denaturing electrophoresis followed by silver staining resulted in distinct banding patterns. Samples possessing a single allele in each amplification group showed two‐band patterns which correspond to the sense and antisense strands, while heterozygotes in the same group or a mixture of two single‐type samples showed four‐band patterns. All of the analysed alleles were discriminated in each DRB1 group. The method described here may be somewhat complicated for routine typing of HLA‐DRB1 alleles. However, it is useful in the screening of ‘new’ alleles as well as the donor‐recipient molecular matching of HLA class II genes for various purposes, e.g. selection of bone marrow transplant donors.