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Updated: Jan 13, 2026
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Dual-modality Molecular Cartography: Integrating Multiplex mRNA Detection with Protein Imaging Mass Cytometry
Published on: November 14, 2025
267
Comprehensive characterization of multivalent mRNA vaccines functionality by flow cytometry and mass spectrometry.
Thibaut Willemin1, Fabien Guinchard1, Jauris Marmey2
1Sanofi Pasteur, Marcy l'Étoile, France.
Vaccine
|January 8, 2026
Summary
Analyzing messenger RNA (mRNA) vaccines requires adaptable methods. Integrated median fluorescence intensity (iMFI) offers a better view of in vitro functionality than cell percentages, while liquid chromatography-mass spectrometry (LCMS) provides an antibody-free quantification alternative.
Area of Science:
- Biotechnology
- Vaccinology
- Analytical Chemistry
Background:
- Clinical trials for mRNA vaccines are rapidly increasing, necessitating robust in vitro analysis methods.
- Current antibody-based methods for assessing mRNA vaccine biological activity have limitations in quantifying antigen expression and platform applicability.
- The complex mechanism of mRNA vaccines demands analytical techniques that are both adaptable and provide accurate quantification.
Purpose of the Study:
- To identify the most relevant readout for antibody-based methods assessing mRNA vaccine biological activity.
- To compare antibody-based methods with an antibody-independent method, liquid chromatography coupled with mass spectrometry (LCMS).
- To evaluate the suitability of these methods for analyzing multivalent mRNA samples in stability studies.
Main Methods:
- Utilized antibody-based methods measuring percentage of positive cells and integrated median fluorescence intensity (iMFI).
- Employed liquid chromatography coupled with mass spectrometry (LCMS) for antibody-independent quantification of antigen expression.
- Analyzed multivalent mRNA samples under stability conditions using both approaches.
Main Results:
- Percentage of positive cells failed to detect differences in protein expression between samples.
- Integrated median fluorescence intensity (iMFI) effectively highlighted differences in protein expression, aligning with LCMS quantification.
- LCMS proved to be a viable antibody-free method for quantifying mRNA vaccine functionality, but cannot perform relative potency analysis due to potential loss of parallelism.
Conclusions:
- Integrated median fluorescence intensity (iMFI) provides a more comprehensive assessment of in vitro mRNA vaccine functionality compared to the percentage of positive cells.
- Liquid chromatography-mass spectrometry (LCMS) is a viable antibody-free alternative for quantifying mRNA vaccine functionality.
- Neither iMFI nor LCMS, as applied in this study, allows for relative potency calculations according to US pharmacopeia guidelines.
