HLA-B27 by PCR
I. INTENDED USE
HLA-B27 Gene Detection Assay provides materials for the isolation of DNA from human whole blood, the in vitro amplification of HLA-B gene sequences, and the subsequent detection of HLA-B27 by group-specific hybridization in microwells.
Human Leukocyte Antigen (HLA) class I specificity B27 represents a family of several closely related alleles (designated B*2701,B*2702, . . .), differing only by a limited number of nucleotide substitutions within exons 2 and 3. The various subtypes also differ in their ethnic distributions, with B*2705 being the most widespread form.
A group of inflammatory rheumatic disorders (collectively termed seronegative spondylo-arthropathies), among them ankylosing spondilitis (Morbus Becherew), Morbus Reiter, and psoriatic arthritis, is known to be highly associated with the presence of the HLA-B27 antigen. Although the exact mechanism determining disease susceptibility is still unknown, testing for HLA-B27 provides a valuable aid for the definite diagnosis of these rheumatic disorders.
III. PRINCIPLES OF THE ASSAY
The HLA-B27 Gene Detection Assay is based on the reverse hybridization principle, and includes three successive steps: DNA is isolated from anticoagulated blood by a rapid and convenient procedure. Then, HLA-B gene (exon 2) sequences are in vitro amplified and terminally labeled with a reporter molecule. Finally, the amplification product is hybridized to a group-specific HLA-B27 (B27) and a generic control (CTL) oligonucleotide probe in two separate cavities of a microwell plate. Bound sequences are detected using a horseradish peroxidase-labeled antibody to the reporter molecule and color substrates.
*Run out time: 48 hours.