HLA TYPING FOR DR3 AND DR4 USING ARTIFICIAL RESTRICTION-FRAGMENT-LENGTH-POLYMORPHISM PCR FROM ARCHIVAL DNA

Aim - To develop polymerase chain reaction based artificial restrictio n fragment length polymorphism (artificial RFLP PCR) assays for DR3 an d DR4 alleles of the multiallelic DRB1 locus and to apply them to para ffin wax embedded archival material. Methods - Sixty five samples from DRB1 typed cell lines were analysed using the artificial RFLP PCR met hod to determine the specificity and sensitivity of the system. Result s - The artificial RFLP method for typing the DRB1 showed 100% accurac y in the 65 samples previously typed using allele specific PCR and ser ology. The samples included 18 combinations of alleles that included D R3, 18 that included DR4, four that were DR3/DR4 heterozygotes, and 10 samples that were neither DR3 nor DR4. Typing of 10 paraffin wax embe dded samples using artificial RFLP PCR was in complete agreement with previous typing at the DRB1 locus. Conclusion - The application of art ificial RFLP PCR for the analysis of multiallelic loci, such as those of the HLA system, in archival DNA samples achieved. Artificial RFLP r obust, easily implemented, non-isotopic system and may be useful for l arge retrospective studies.