SEROLOGICAL VERSUS MOLECULAR CHARACTERIZATION OF CLASS II HLA FOR RECIPIENTS AND DONORS OF BONE MARROW TRANSPLANTATION

Abstract

The choice of a properly matched donor is the first most important task in bone marrow transplantation. HLA class II ORB* I proved to be the most important factor in determining the degree of graft rejection and graft versus host disease. One hundred and thirty five cases were subjected to different ORB* I typing techniques of varying levels of resolution. Comparative serological versus molecular typing was available for 35 cases , where 20% of results were discordant This is due to cross reactivity and lack of sensitivity and specificity of the serological method . PCR-SSOP was applied using 2 comparative kits, INNO-LiPA kit (32 probes) was used for cases while Amplicore Roche kit (20 probes) was used for 67 cases. lNNO-LiPA gave better resolution at both the haplotype and allele levels. Using SSOP (JNNO-LiPA) commercial kit, single allele (high resolution) could be identified in 28 occasions in certain haplotype combinations and in ORB1*070 I and *I 00 I being single allele. Single allele was identified in 56 other haplotypes representative of most ORB I specificities starting by INNO-LiPA followed by a modified PCR-RFLP making advantage of the commercial kit complemented by a limited number of specific primers and restriction enzymes. For class II, serological typing is not anymore the method of choice; ORB! has to be molecularly characterized. Commercially available kits depending on SSOP give an intermediate level of resolution, to reach a high level a complementary modified PCR-RFLP was introduced. With the final aim of applying this technique to different donor selection protocols, whether for siblings, family members, unrelated donors or cord blood. This is a practical economic and simple technique which could be a reasonable substitute for the sequence-based typing when it is not available, however, it can not detect new described alleles.