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Visualizing Antigen Specific CD4+ T Cells using MHC Class II Tetramers
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Techniques used to define human MHC antigens: serology

P A Dyer1, S Martin

  • 1Northwestern Regional Tissue Typing Laboratory, St. Mary's Hospital, Manchester, U.K.

Immunology Letters
|July 1, 1991
PubMed

Insights

The complement-mediated cytotoxicity assay remains the standard for human leukocyte antigen (HLA) tissue typing, defining HLA antigens on various cell types. Automation and microsphere technology enhance this established method for accurate HLA typing.

Area of Science:

  • Immunology
  • Genetics
  • Clinical Diagnostics

Background:

  • The complement-mediated cytotoxicity assay, developed in the 1960s, is the classical method for human leukocyte antigen (HLA) tissue typing.
  • This assay defines HLA antigens on lymphocytes from peripheral blood, spleen, or lymph nodes, crucial for transplantation and disease studies.

Purpose of the Study:

  • To describe the established complement-mediated cytotoxicity assay for HLA antigen definition.
  • To highlight the application of this assay in various clinical and research settings, including cell lines and prenatal diagnostics.

Main Methods:

  • Target cells (lymphocytes) are incubated with typing antisera, followed by the addition of rabbit serum as a complement source.
  • Incubation at 22°C in Terasaki trays, followed by staining to detect target cell death, indicates a positive reaction.
  • Automation and antibody-coated magnetisable microspheres are employed for enhanced efficiency and pure B lymphocyte separation for class II typing.

Main Results:

  • The assay accurately defines HLA-A, B, Cw (class I) and HLA-DR, DQ (class II) antigens.
  • Successful HLA typing has been demonstrated using various cell sources, including cultured cells and cell lines.
  • The method allows for the definition of antibodies to HLA antigens using cells with known HLA phenotypes.

Conclusions:

  • The complement-mediated cytotoxicity assay is a robust and adaptable method for HLA antigen definition.
  • Modern adaptations, including automation and microsphere technology, improve the efficiency and precision of HLA typing.
  • This assay remains fundamental for immunological and clinical applications requiring precise HLA characterization.

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