High-Throughput Quantification of Spike Protein Expression by LC-MS for mRNA-LNP Evaluation and Immune Response...

Area of Science:

• Biochemistry
• Immunology
• Vaccinology

Background:

• The COVID-19 pandemic increased interest in immuno-multiple reaction monitoring (immuno-MRM) for peptide quantification.
• The rise of mRNA vaccines necessitates robust methods for quantifying antigen expression.

Purpose of the Study:

• To present an optimized immuno-MRM method for quantifying SARS-CoV-2 spike fusion peptide expression after mRNA or plasmid transfection.
• To enable accurate assessment of protein translation in vitro and analyze host cell proteomic responses.

Main Methods:

• Developed and validated an immuno-MRM assay targeting the SARS-CoV-2 spike fusion peptide SFIEDLLFNK.
• Utilized a dual-function workflow to measure antigen levels and analyze flow-through for proteomic insights.

Main Results:

• The optimized immuno-MRM method demonstrated high sensitivity, precision, and linearity across a wide dynamic range.
• The assay accurately quantifies protein translation in vitro.
• Flow-through analysis provided insights into host cell proteomic responses.

Conclusions:

• The developed immuno-MRM method is a sensitive and precise tool for quantifying antigen expression from mRNA vaccines.
• This workflow supports comprehensive characterization of mRNA vaccine efficacy and safety.
• The method is valuable for vaccine development, quality control, and regulatory evaluation of RNA-based therapeutics.