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Report from the HLA class II typing by PCR-SSP Multicentre Study
H Zetterquist1, M Bengtsson1, G Bäckström1
1Centre for BioTechnology, Karolinska Institute, NOVUM, Huddinge and Department of Clinical Immunology, Karolinska Institute at Huddinge Hospital, Huddinge, Sweden; ,Department of Clinical Immunology, Academic Hospital, Uppsala, Sweden; ,Department of Forensic Medicine, University Hospital, Linköping, Sweden; ,Regional Centre for Blood Transfusion and Clinical Immunology, Aalborg Hospital, Aalborg, Denmark; ,Regional Blood Centre, Sahlgren's Hospital, Gothenburg, Sweden; ,Institute of Transplantation Immunology, National Hospital, Oslo, Norway; ,Department of Clinical Immunology, University Hospital of Aarhus, Aarhus, Denmark; ,Department of Clinical Immunology, National Hospital, Copenhagen, Denmark; and ,Regional Blood Centre, University Hospital, Lund, Sweden.
This study demonstrates the utility of Polymerase Chain Reaction Sequence-Specific Primers (PCR-SSP) for rapid HLA-DR and HLA-DQ typing, achieving high accuracy despite minor amplification failures. The method efficiently defines key specificities in under 2.5 hours.
Area of Science:
- Immunogenetics
- Molecular Biology
- Clinical Diagnostics
Background:
- Human Leukocyte Antigen (HLA) typing is crucial for transplantation and disease association studies.
- Low-resolution HLA-DR and HLA-DQ typing provides essential information for clinical applications.
- Rapid and accurate typing methods are needed to meet clinical demands.
Purpose of the Study:
- To evaluate the utility of Polymerase Chain Reaction Sequence-Specific Primers (PCR-SSP) for routine, low-resolution HLA-DR and HLA-DQ typing.
- To assess the turnaround time and accuracy of the PCR-SSP method across multiple laboratories.
- To identify potential challenges and optimize the protocol for high-throughput HLA typing.
Main Methods:
- Utilized PCR-SSP for low-resolution HLA-DR and HLA-DQ typing across nine laboratories.
- Employed a rapid bromide salt DNA extraction protocol.
- Analyzed results from 360 typings using pre-aliquoted PCR trays for efficiency.
Main Results:
- Achieved 98% accuracy for HLA-DR typings and 89% for HLA-DQ typings compared to reference typing.
- Typing, including DNA extraction, was completed within 2.0 to 2.3 hours.
- Observed a 3% PCR amplification failure rate, impacting 52% of typings, but maintained a 91-98% success rate for correctly identified alleles.
Conclusions:
- PCR-SSP is a valuable tool for rapid, routine low-resolution HLA-DR and HLA-DQ typing.
- The method demonstrates high accuracy and efficiency, suitable for clinical settings.
- Despite amplification failures, the overall success rate for allele identification remains high, indicating robustness.