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TaqMan probe real-time PCR for quantitative detection of bovine adenovirus type 1 during the manufacture of biologics and medical devices using bovine-derived raw materials

소유래 성분 원재료 사용 생물의약품과 의료기기 제조 공정에서 bovine adenovirus type 1 정량 검출을 위한 TaqMan probe real-time PCR

  • Ko, Woon Young (Center for Biopharmaceuticals Safety Validation, BioPS Co., Ltd. Daedeuk Valley Campus, Hannam University) ;
  • Noh, Na Gyeong (Department of Biological Sciences and Biotechnology, Hannam University) ;
  • Kim, In Seop (Department of Biological Sciences and Biotechnology, Hannam University)
  • 고운영 ((주)바이오피에스 바이오의약품안전성검증센터) ;
  • 노나경 (한남대학교 생명.나노과학대학 생명시스템과학과) ;
  • 김인섭 (한남대학교 생명.나노과학대학 생명시스템과학과)
  • Received : 2015.08.19
  • Accepted : 2015.09.11
  • Published : 2015.09.30

Abstract

Biologics and medical devices manufactured with bovine-derived raw materials have the risk of viral contamination. Therefore, viral validation study is essential to ensure the safety of the products. Bovine adenovirus type-1 (BAdV-1) is one of the common bovine viral pathogens. For quantitative detection of BAdV-1 during the manufacture of biologics and medical devices, a TaqMan probe real-time PCR method was developed. Specific primers and TaqMan probe for amplifying and detecting BAdV-1 DNA were designed. Specificity, limit of detection (LOD), and robustness of the method was validated according to international guideline on the validation of nucleic acid amplification tests for the pathogen detection. The sensitivity of the assay was found to be $7.44{\times}10^1\;TCID_{50}/ml$. The real-time PCR method was reproducible, very specific to BAdV-1, and robust. Moreover, the method was successfully applied to the validation of Chinese Hamster Ovary (CHO)-K1 cells artificially infected with BAdV-1, a commercial CHO master bank, and bovine type 1 collagen. The overall results indicate that this rapid, specific, sensitive, and robust assay can be reliably used for quantitative detection of BAdV-1 contamination during the manufacture of biologics and medical devices using bovine-derived raw materials.

Keywords

biologics;bovine adenovirus;medical device;TaqMan probe real-time PCR;virus detection

Acknowledgement

Supported by : 한국연구재단

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