Systematic analysis of COVID-19 mRNA vaccines using four orthogonal approaches demonstrates no excessive DNA impurities

Robust epidemiologic data confirm that vaccination saved millions of lives and protected recipients from severe COVID-19 disease caused by SARS-CoV-2 infection. Despite this evidence, COVID-19 mRNA vaccines have become a target of largely unfounded skepticism built on coincidental health events occurring after vaccination without proven causality. An example of claims undermining trust in mRNA vaccine safety relates to excessive amounts of residual DNA from the vaccine manufacturing process. These claims are based on improperly accomplished quantification methods and/or incorrectly interpreted data. In this study, we aimed to thoroughly assess quantity, quality and identity of residual DNA in mRNA vaccines based on objective, technically correctly performed and interpreted methods using orthogonal approaches, including qPCR, fluorometry, capillary electrophoresis and short-read DNA sequencing. We analysed 15 batches of Comirnaty and Spikevax vaccines available in Slovakia, obtained from official repositories under the supervision of the State Institute for Drug Control. These included 11 batches post-expiration date that were previously claimed by others to contain “significant amounts of DNA” using data from reverse transcription-based multiplex qPCR performed with technical shortcomings. Results of our analyses utilising four orthogonal approaches clearly demonstrate that the quantity of residual DNA in all analysed vaccine batches is below approved limits, in very low quantities relative to mRNA, and degraded into small fragments, which originate from the DNA template used to transcribe mRNA during vaccine production. Our results show that reliable vaccine analysis requires rigorous application of multiple validated methods that comply with molecular characteristics of mRNA vaccine components and minimise their mutual interference.