A cell-based multiplex immunoassay platform using fluorescent protein-barcoded reporter cell lines.

Area of Science:

• Biotechnology
• Immunology
• Cell Biology

Background:

• Multiplex immunoassays commonly use acellular antigens on solid-phase platforms like Luminex technology.
• Cell barcoding with fluorescent dyes allows multiplex cell-based assays but involves complex labeling and quality control steps.
• A need exists for a more streamlined approach to multiplex screening of antibodies targeting membrane proteins.

Purpose of the Study:

• To develop and validate a novel cell-based platform using stable, fluorescent protein-barcoded reporter cell lines for multiplex antibody screening.
• To enable high-throughput characterization of immune responses and discovery of antibodies against membrane-associated proteins.

Main Methods:

• Generation of stable reporter cell lines engineered with fluorescent proteins for multiplexing.
• Utilizing flow cytometry for deconvolution of barcoded cell panels.
• Testing the system with cell panels expressing influenza A hemagglutinin trimers, human CCR2, and CCR5 proteins, including epitope-defined mutants.

Main Results:

• Successful generation of stable, fluorescent protein-barcoded reporter cell lines.
• Demonstrated utility of the cell-based system for multiplex screening via flow cytometry deconvolution.
• Validated the platform with diverse membrane proteins, including viral and human targets, and their mutants.

Conclusions:

• The developed fluorescent protein-barcoded reporter cell line system offers a robust and efficient platform for multiplex antibody screening.
• This technology has the potential for 256-plex cell panels, significantly advancing antibody discovery and immune profiling for membrane proteins.