Development of a simple, fast, and accurate HLA-DNA typing method in the organ transplantation

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HLA typing of both donor and recipients in organ transplantation is important from the stand point of equity and efficiency. In this research, we have aimed at developing a simple, fast, and accurate HLA-DNA typing system, a chemiluminescence-based Hydridization Protection Assay (HPA) technology, to be used for the organ transplantation. RNA was amplified by a method of Transcription-Mediated Amplification (TMA) targeting at the genomic DNA and was detected by HPA which is one type of sequence-specific oligonucleotide (SSO) methods. Twenty-four probes were used to detect 21 HLA-DRB1 alleles or allele groups in 28 HLA-homozygous cell lines and 158 normal volunteers. The results were typable in 99.4% of the samples and completely concordant with the serological typing. DR1, DR7, DR9, DR11, DR12, DR13, DR1403, and HR5 were perfectly matched with the serology by one-to-one correspondence. DR2, DR4, DR8, and DR14 were typed in more detail by TMA-HPA.Operating time required for extraction of DNA was one hour, for amplification 4 to 5 hours, for detection one hour, a total of 6 to 7 hours. We further typed HLA-DRB1 alleles of 97 patients in the waiting list for renal transplantation in Saitama prefecture and obtained reliable results, which indicated the data conversion in both ways between serology and DNA typing are possible. Retyping of unreliable serologic DR typings by TMA-HPA was recommended to assure the equity and efficiency in organ transplantation.