HLA-DR Typing by Polymerase Chain Reaction with Sequence- Specific Primers Compared to Serological typing

Abstract

Background: Considering the role of HLA matching in transplant outcome, the quality of HLA-DR typing is clearly an important issue. In recent years, serological methods have been replaced with DNA based typing methods. The main objective of this study was to compare HLA-DR typing data obtained from existing serologic method with data obtained by the new PCR-SSP method. Methods: 55 peripheral blood samples were collected from randomly selected individuals who were referred to the transplantation laboratory of Isfahan, in Aliasghar Hospital, and were typed for HLA-DR antigens by both methods. HLA-DR typing by serologic method was performed using 30 different antisera and for PCR-SSP method, specific primers were used for HLA-DRB1*01-10(except DR6, 8, 10), and also for HLA-DR52, and DR53. After DNA extraction, 13 pairs specific primers were used for each sample separately and PCR reaction were done. In this study, the third intron of DR locus was used as internal positive control. After PCR amplification, products of reaction electrophoresis was performed on 2% agarose gel, and after taking photo of gel, interpretation and comparison of results were performed. Results: The results of 31 samples (56.3%) corresponded completely to serological method, 12 samples (22%) were assigned heterozygous in serology and homozygous in molecular typing, 7 samples (12.7%) were heterozygous in both methods but different in one allele. 2 samples (3.6%) were homozygous in serology and heterozygous in molecular typing, and also one sample (1.8%) was homozygous in both methods but so that in serology DR14, and in molecular typing DR11 were assigned. And finally 2 samples from 55 (3.6%) were not detectable in serological method. Conclusion: The typing data obtained from the conventional and the new methods were compared. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were calculated. The results indicated that the DNA based method had more sensitivity, accuracy, and resolving power than serologic typing methods.
Keywords: HLA-DR, PCR-SSP, serological typing.