Reference strand conformation analysis for determination of HLA polymorphisms

The methods used for analysis of HLA gene polymorphisms differ according to their resolution capacity. We compared the resolution of Reference Strand Conformation Analysis (RSCA) with that of three other methods, i.e. Sequence Specific Oligonucleotide Probes (SSOP), Sequence Specific Primers (SSP) and Sequence Based Typing (SBT). Ninety-six healthy individuals were typed. Fifty-four individuals were typed for HLA-A and -B loci at low resolution (LR) - by SSP and SSOP - and at high resolution level (HR) by SSP and RSCA. The other 44 individuals were typed for DRB1 and DQB1 loci at LR level and as well at HR level by SSP and SBT. Polymorphisms in exon 2 of HLA class II genes and in exons 2 and 3 of class I genes were defined. We found a 100% concordance between the SSP and SSOP typing results at the LR level of all (A, B DRB1 and DQB1) loci. HR typing of class I by RSCA revealed 87% concordance with the SSP method for locus A and 100% concordance for locus B. HR typing of class II by SBT reached the 100% concordance with SSP results in all tested loci. Conclusion: To our needs the RSCA method was not successful in HR typing, while by combining the SSP and the SBT system we are able to distinguish a vast majority of HLA alleles.