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Updated: Oct 12, 2025
Genetic Barcoding with Fluorescent Proteins for Multiplexed Applications
Published on: April 14, 2015
A cell-based multiplex immunoassay platform using fluorescent protein-barcoded reporter cell lines
Shengli Song1, Miriam Manook2, Jean Kwun2
1Department of Immunology, Duke University School of Medicine, Durham, North Carolina, USA.
Insights
Researchers developed stable, fluorescent protein-barcoded reporter cell lines for multiplex antibody screening against membrane proteins. This new cell-based platform enables high-throughput antibody discovery and immune characterization.
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.
Abstract:
Multiplex immunoassays with acellular antigens are well-established based on solid-phase platforms such as the Luminex® technology. Cell barcoding by amine-reactive fluorescent dyes enables analogous cell-based multiplex assays, but requires multiple labeling reactions and quality checks prior to every assay. Here we describe generation of stable, fluorescent protein-barcoded reporter cell lines suitable for multiplex screening of antibody to membrane proteins. The utility of this cell-based system, with the potential of a 256-plex cell panel, is demonstrated by flow cytometry deconvolution of barcoded cell panels expressing influenza A hemagglutinin trimers, or native human CCR2 or CCR5 multi-span proteins and their epitope-defining mutants. This platform will prove useful for characterizing immunity and discovering antibodies to membrane-associated proteins.
