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Related Experiment Videos

Automated HLA-B27 testing using the FACSPrep/FACScan system

W M Reynolds1, P R Evans, A C Lane

  • 1Wessex Histocompatibility Group, Southampton University Hospitals NHS Trust, Shirley, United Kingdom.

Cytometry
|June 15, 1994
PubMed
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A new flow cytometry method enables rapid, large-scale screening for HLA-B27, a marker for ankylosing spondylitis. While effective for negative screening, cross-reactivity with HLA-B7 requires confirmatory tests for definitive HLA-B27 typing.

Area of Science:

  • Immunogenetics
  • Clinical Diagnostics
  • Flow Cytometry

Background:

  • Ankylosing spondylitis, a seronegative arthropathy, is often associated with the HLA-B27 tissue type.
  • Accurate and efficient HLA-B27 screening is crucial for diagnosis and patient management.
  • Existing methods may lack the speed and throughput for large-scale screening.

Purpose of the Study:

  • To develop and evaluate a rapid, reproducible flow cytometry-based assay for HLA-B27 screening using whole blood.
  • To assess the feasibility of large-scale sample screening with an automated system.
  • To identify limitations and potential improvements for routine HLA-B27 typing.

Main Methods:

  • Utilized an automated sample preparation system (FACSPrep/FACScan) for flow cytometry analysis.

Related Experiment Videos

  • Employed both indirect and direct monoclonal antibody staining techniques with whole blood samples.
  • Assessed results using median channel shift (CS) and relative fluorescence intensity, analyzing cross-reactivity with HLA-B7.

    • Main Results:

    • The developed flow cytometry assay demonstrated rapidity, reproducibility, and capacity for simultaneous screening of large sample numbers.
    • Indirect staining with HLA-ABC-m3 showed significant CS differences between HLA-B27 and HLA-B7 in 60% of cases.
    • Direct dual staining allowed for effective negative screening, eliminating over 60% of samples (CS < 15) and differentiating HLA-B7 and HLA-B27 based on CS values.

    Conclusions:

    • The flow cytometry method provides a valuable tool for rapid, large-scale negative screening of HLA-B27.
    • Monoclonal antibody cross-reactivity with HLA-B7 necessitates confirmatory testing for definitive HLA-B27 identification.
    • Using an HLA-B27 specific monoclonal antibody (FD705) could enhance the assay's utility for routine typing.