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KMID : 0364820150510030199
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Korean Journal of Microbiology
2015 Volume.51 No. 3 p.199 ~ p.208
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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
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Ko Woon-Young
Noh Na-Gyeong
Kim In-Seop
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Abstract
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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 × 101 TCID50/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, acommercial 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.
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KEYWORD
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biologics, bovine adenovirus, medical device, TaqMan probe real-time PCR, virus detection
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