Rapid Analysis of mRNA 5’ Capping and Poly-A Tailing with High-Resolution LC-MS

Industrial-scale production of mRNA has taken on renewed importance due to the on-going pandemic. Efficient translation of mRNA to protein depends critically on: (i) 5’-terminal capping, and (ii) 3’ terminal polyadenylation, which can be considered critical quality attributes that must be characterized and monitored.

This presentation will show that:

• mRNA capping can be analyzed by LC-MS within 75 minutes by taking advantage of a thermostable enzyme to rapidly liberate 5’ terminal oligonucleotides.
• Certain enzymes commonly used during in vitro transcription can incorporate non-templated nucleotides into both 5’ and 3’ terminal regions.
• Non-templated nucleotides can be considered product-related impurities and can be detected and quantified by LC-MS.

Three learning objectives:

• LC-MS is a robust, sensitive and specific technology for quality control of mRNA.
• Workflows for analysis of mRNA 5’ capping and poly-A tailing
• Detection and characterization of product-related impurities stemming from incorporation of non-templated nucleotides

Presenter: Brian Liau, PhD (Solution Development Scientist, Singapore, Agilent Technologies, Inc.)

Brian Liau obtained his bachelor’s degree at Johns Hopkins University and his PhD at Duke University. Currently based at Agilent’s Global Solution Development Center in Singapore, he develops new LC-MS and spectroscopic techniques to ensure the quality and safety of next-generation therapies such as AAV vectors and mRNA vaccines. Prior to joining Agilent, he was a research fellow at the Bioprocessing Technology Institute (A*STAR).