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Related Experiment Videos

HLA-B27 screening by flow cytometry

B Lingenfelter1, T C Fuller, L Hartung

  • 1Department of Pathology, University of Utah, Salt Lake City, USA.

Cytometry
|June 15, 1995
PubMed
Summary
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A new flow cytometry method accurately detects the lymphocyte Human Leukocyte Antigen B27 (HLA-B27) expression. This convenient assay shows high sensitivity and specificity for screening HLA-B27, aiding in clinical diagnosis.

Area of Science:

  • Immunology
  • Clinical Diagnostics
  • Flow Cytometry

Background:

  • Human Leukocyte Antigen B27 (HLA-B27) is a crucial marker in certain autoimmune diseases.
  • Traditional microlymphocytotoxicity testing for HLA-B27 is time-consuming and complex.
  • Development of rapid and accurate methods for HLA-B27 detection is clinically significant.

Purpose of the Study:

  • To evaluate a novel two-color direct immunofluorescent flow cytometric assay for lymphocyte HLA-B27 expression.
  • To compare the performance of the flow cytometric assay against traditional microlymphocytotoxicity testing.
  • To determine the sensitivity and specificity of the flow cytometric HLA-B27 assay.

Main Methods:

  • A two-color direct immunofluorescent assay using fluorescein-isothiocyanate (FITC)-conjugated anti-B27 and phycoerythrin (PE)-conjugated anti-CD3 antibodies.

Related Experiment Videos

  • Whole blood samples from 209 clinical cases were analyzed using flow cytometry.
  • Analysis involved gating on CD3 positive events and evaluating anti-B27 staining intensity via median channel fluorescence.

    • Main Results:

    • The flow cytometric assay demonstrated high sensitivity (97.6%) and specificity (95.9%) for HLA-B27 detection.
    • Median channel fluorescence correlated with HLA-B27 expression levels, distinguishing negative, B7 positive, and B27 positive samples.
    • Cross-reactivity was observed with HLA-B7 (n=38), HLA-B37 (n=3), and HLA-B39 (n=3) antigens.

    Conclusions:

    • The flow cytometric assay for lymphocyte HLA-B27 expression is a convenient and useful screening tool.
    • The assay offers high sensitivity and specificity, comparable to conventional methods.
    • Confirmation with microlymphocytotoxicity or alternative monoclonal antibodies is recommended for positive flow cytometry results to maximize specificity.