HLA typing

The OUHSC Laboratory for Genomics and Bioinformatics currently offers HLA typing using methods developed by Dr. William Hildebrand of the Department of Microbiology and Immunology. The human HLA class I and class II molecules mediate many important immune functions. These molecules trigger anti-bacterial and anti-viral immune responses and are therefore important for vaccine development and for understanding susceptibility and resistance to infection. Class I and class II molecules also mediate autoimmune responses such as diabetes and arthritis. These molecules must be matched for the transplantation of organs (kidney, heart, liver) and bone marrow. Each individual inherits a different set of class I and class II molecules. We all therefore differ in how we respond to infection, vaccination, and from whom we should receive a transplant. Indeed, HLA typing is primarily utilized for the clinical matching of transplant recipients and donors.

Today, the gold standard in matching class I and class II HLA molecules for transplantation is by DNA sequencing. The Hildebrand laboratory pioneered this technology. Because this sequence-based HLA typing can be done in a cost-effective and robust manner, HLA class I and class II DNA sequence-based typing (SBT) also has become the norm for research studies. Consequently, Dr. Hildebrand's laboratory has completed nearly 20,000 HLA typings in the past 8 years as part of their cancer research studies.

High resolution sequence-based typing is performed using a solid phase technology. Genomic DNA is extracted from whole blood or cells and class I or class II specific PCR is then performed. For class I molecules all of exon 2 and exon 3 are sequenced in both directions. For class II molecules all of exon 2 is sequenced. This precisely establishes the HLA type of an individual. In addition to being an established research protocol, Dr. Hildebrand's laboratory is CLIA/ASHI accredited, and can provide transplant patients in Oklahoma with the highest quality HLA type for transplantation.

For more information or ordering please contact william-hildebrand@ouhsc.edu

 

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Figure 1. Sequence-based-typing strategy for direct heterozygous sequencing of HLA-A,B exons 2 and 3. Genomic DNA, represented here as a linear diagram of exons 2 and 3 flanked by introns 1, 2, and 3, acts as template for the primary (1o) PCR reaction. HLA-A, B locus specific oligonucleotide primers (5'AI1 and 3'AI3 for HLA-A) hybridize in introns 1 and 3 resulting in a primary amplicon of approximately 914 bp in length. This primary locus-specific PCR product then serves as template for four nested and hemi-nested secondary (2o) PCR reactions, shown below the linear diagram of exons 2 and 3. Oligonucleotide primer mixes A2, A3, A4, and A5 generate separate exon 2 and 3 HLA-A amplicons of approximately 340 bp in length with an M13 universal primer site on one end and biotinylated on the other. The HLA-A, -B exon 2 and 3 biotinylated PCR products are then bound to a streptavidin-coated support upon which bi-directional DNA sequencing reactions are performed.