[Rapid genotyping for HLA-DR by PCR-amplification with sequence-specific primers and clinical practice]

Genotyping for HLA-DR alleles by polymerase chain reaction with sequence-specific primers (PCR-SSP) was first typed in 112 individuals of donor-recipients of cadaveric transplantation and 9 cell lines DNA. Twenty separate PCR reactions were performed per sample. The amplification was accomplished by 34 cycles consisting of denaturation at 94 degrees C for 30 sec, annealing at 60 degrees C for 50 sec and extension at 72 degrees C for 40 sec. HLA-DR alleles could be accurately distinguished. The overall time of genotyping was only 5 hours. The specificity of matching was determined against a panel of standard DNA, analysis with restriction endonucleases, and Southern hybridization using DIG oligonucleotide 3'- end labeling probes. The specificity and reproducibility were 100%. No false positive or false negative typing results were obtained. These showed genotyping by PCR-SSP was a rapid and accurate matching technique, suited for clinical practice.