HIGH-RESOLUTION HLA-DPB TYPING BASED UPON COMPUTERIZED ANALYSIS OF DATA OBTAINED BY FLUORESCENT SEQUENCING OF THE AMPLIFIED POLYMORPHIC EXON-2

To differentiate 32 HLA-DPB alleles, conventional techniques such as s erology and cellular typing are inadequate for high-resolution DPB typ ing. The most refined DNA typing until now is SSO typing and new selec ted oligonucleotides can be added to this system to distinguish new al lele sequences. DNA sequencing, however, reveals directly the sequence information of all polymorphic HVRs and has the advantage of being in dependent from exon polymorphisms. We have developed a new DNA-based t yping approach that is rapid, fully automated, and therefore suitable for routine typing. The system is based upon direct sequencing of ampl ified DNA with fluorescent-labeled primers. The designation of alleles is obtained by a comparison of all polymorphic positions in the deter mined sequence with all known allele sequences retained in a database along with their heterozygous combinations. Sequence data at both cons tant and polymorphic positions are used for quality control. In this s tudy, the typing results of a panel of 91 previous SSO-typed DNA sampl es are described. After comparison with the SSO-typing results, we con clude that with this SBT system allele assignment is reliable. The met hod is easy to perform since both sequencing and assignment are automa ted. Furthermore, the system is easily applicable to other gene system s.