Three analytical methods could track , with different strengths for resolution, throughput, and detecting functional oligomers.
Evidence
This comparative analytical-methods study evaluated capillary gel electrophoresis with fluorescent detection, capillary gel electrophoresis with UV detection, and IP-RPLC with UV detection on mRNA drug substance and drug product samples.
Caveat
The study characterizes analytical performance and protein expression from oligomers, not clinical vaccine efficacy or long-term product stability.
Simplified
UNLABELLED: Development of mRNA technologies has opened new horizons for both preventive vaccines and therapeutics. The rapid growth of mRNA products goes hand in hand with the need for novel analytical methods to characterize these products. One common critical quality attribute of mRNA Drug Substance (DS) and Drug Product (DP) is . mRNA molecules are susceptible to degradation, often necessitating ultra-low-temperature storage of mRNA vaccines. Once degraded, mRNA can no longer be correctly translated into the target protein, leading to reduced vaccine efficacy. Therefore, robust analytical methods are crucial for understanding mRNA degradation mechanisms, optimizing formulation stability, and maximizing efficacy. In this study, we compared three analytical methods to assess mRNA integrity and purity: capillary gel electrophoresis (CGE) with fluorescent detection using a fragment analyzer (FA) device, CGE with ultraviolet (UV) detection, and ion-pairing reversed-phase chromatography (IP-RPLC) with UV detection. All three methods successfully monitored the integrity of mRNA DS and DP. IP-RPLC provided better resolution and facilitated the detection of , while the FA, being more automated, enabled rapid analysis of a large number of samples. CGE-UV revealed the presence of mRNA oligomers. Further analysis showed that these oligomers could express the target protein similarly to the mRNA monomer.
SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1038/s41598-025-27563-z.
Key numbers
2%
Day 15 in DP
Main peak area of in Drug Product at day 15.
70%
Day 15 in DS
Main peak area of in Drug Substance at day 15.
1.2%
Day-to-Day Variability in
Lower variability in assessment using .
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