Validation of a real-time RT-PCR method to quantify newcastle disease virus (NDV) titer and comparison with other quantifiable methods

Juno Jang, Sung Hwan Hong, Ik Hwan Kim

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

A method for the rapid detection and quantification of Newcastle disease virus (NDV) produced in an animal cell culture-based production system was developed to enhance the speed of the NDV vaccine manufacturing process. A SYBR Green I-based real-time RT-PCR was designed with a conventional, inexpensive RT-PCR kit targeting the F gene of the NDV LaSota strain. The method developed in this study was validated for specificity, accuracy, precision, linearity, limit of detection (LOD), limit of quantification (LOQ), and robustness. The validation results satisfied the predetermined acceptance criteria. The validated method was used to quantify virus samples produced in an animal cell culture-based production system. The method was able to quantify the NDV samples from mid- or late-production phases, but not effective on samples from the early-production phase. For comparison with other quantifiable methods, immunoblotting, plaque assay, and tissue culture infectious dose 50 (TCID50) assay were also performed with the NDV samples. The results demonstrated that the real-time RT-PCR method is suitable for the rapid quantification of virus particles produced in an animal cell-culture-based production system irrespective of viral infectivity.

Original languageEnglish
Pages (from-to)100-108
Number of pages9
JournalJournal of microbiology and biotechnology
Volume21
Issue number1
DOIs
Publication statusPublished - 2011 Jan

Keywords

  • Method comparison
  • Method validation
  • Newcastle disease virus (NDV)
  • Real-time RT-PCR (RRT-PCR)
  • Virus quantification

ASJC Scopus subject areas

  • Biotechnology
  • Applied Microbiology and Biotechnology

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