A Versatile Simple Capture Assay for Assessing the Structural Integrity of MHC Multimer Reagents

Area of Science:

• Immunology and Molecular Biology
• Biotechnology and Reagent Development

Background:

• Major Histocompatibility Complex (MHC):peptide multimers are essential tools for visualizing antigen-specific T cell responses.
• Assessing the structural integrity of MHC multimer reagents is crucial before use, particularly with limited or irreplaceable samples, but robust controls are often unavailable.

Purpose of the Study:

• To develop and validate a method for probing the structural integrity of MHC multimers.
• To enable quality control of MHC multimer reagents prior to their use in critical immunological studies.

Main Methods:

• A capture assay was designed using beads coated with anti-mouse immunoglobulin (Ig).
• These beads were incubated with conformation-specific mouse monoclonal antibodies, followed by fluorescently tagged MHC multimers.
• The capture efficiency of labeled multimers by the beads was measured semi-quantitatively using flow cytometry.

Main Results:

• The developed method allows for the visualization of correct MHC multimer folding.
• It enables semi-quantitative comparison of different batches of MHC multimers for quality control.
• The assay can assess the fidelity of MHC multimer structure, including interactions between heavy chain, peptide, and β2M, depending on antibody availability.

Conclusions:

• This novel capture assay provides a reliable method to assess MHC multimer structural integrity.
• It is particularly valuable for studies involving precious or irreplaceable samples, such as patient samples in clinical trials.
• The approach enhances the reliability of T cell response visualization using MHC:peptide multimers.