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A novel method for quantitation of AAV genome integrity using duplex digital PCR

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  • Lauren Tereshko
  • Xiaohui Zhao
  • Jake Gagnon
  • Tinchi Lin
  • Trevor Ewald
  • Yu Wang
  • Marina Feschenko
  • Cullen Mason

Abstract

Recombinant adeno-associated virus (rAAV) vectors have become a reliable strategy for delivering gene therapies. As rAAV capsid content is known to be heterogeneous, methods for rAAV characterization are critical for assessing the efficacy and safety of drug products. Multiplex digital PCR (dPCR) has emerged as a popular molecular approach for characterizing capsid content due to its high level of throughput, accuracy, and replicability. Despite growing popularity, tools to accurately analyze multiplexed data are scarce. Here, we introduce a novel statistical model to estimate genome integrity from duplex dPCR assays. This work demonstrates that use of a Poisson-multinomial mixture distribution significantly improves the accuracy and quantifiable range of duplex dPCR assays over currently available models.

Suggested Citation

  • Lauren Tereshko & Xiaohui Zhao & Jake Gagnon & Tinchi Lin & Trevor Ewald & Yu Wang & Marina Feschenko & Cullen Mason, 2023. "A novel method for quantitation of AAV genome integrity using duplex digital PCR," PLOS ONE, Public Library of Science, vol. 18(12), pages 1-18, December.
    • Handle: RePEc:plo:pone00:0293277
    DOI: 10.1371/journal.pone.0293277
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    References listed on IDEAS

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    1. Simant Dube & Jian Qin & Ramesh Ramakrishnan, 2008. "Mathematical Analysis of Copy Number Variation in a DNA Sample Using Digital PCR on a Nanofluidic Device," PLOS ONE, Public Library of Science, vol. 3(8), pages 1-9, August.
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