Universal real-time PCR assay for quantitation and size evaluation of residual cell DNA in human viral vaccines.

Autor: André M; Agence Nationale de Sécurité du Médicament et des Produits de Santé (ANSM), Control Directorate, Biological Controls for Immunological Medicines and Biological Safety Department, 321 Avenue Jean Jaurès, F-69007, Lyon, France. Electronic address: murielle.andre@ansm.sante.fr., Reghin S; Agence Nationale de Sécurité du Médicament et des Produits de Santé (ANSM), Control Directorate, Biological Controls for Immunological Medicines and Biological Safety Department, 321 Avenue Jean Jaurès, F-69007, Lyon, France., Boussard E; Agence Nationale de Sécurité du Médicament et des Produits de Santé (ANSM), Control Directorate, Biological Controls for Immunological Medicines and Biological Safety Department, 321 Avenue Jean Jaurès, F-69007, Lyon, France., Lempereur L; Agence Nationale de Sécurité du Médicament et des Produits de Santé (ANSM), Control Directorate, Biological Controls for Immunological Medicines and Biological Safety Department, 321 Avenue Jean Jaurès, F-69007, Lyon, France., Maisonneuve S; Agence Nationale de Sécurité du Médicament et des Produits de Santé (ANSM), Control Directorate, Biological Controls for Immunological Medicines and Biological Safety Department, 321 Avenue Jean Jaurès, F-69007, Lyon, France.
Jazyk: angličtina
Zdroj: Biologicals : journal of the International Association of Biological Standardization [Biologicals] 2016 May; Vol. 44 (3), pp. 139-49. Date of Electronic Publication: 2016 Mar 28.
DOI: 10.1016/j.biologicals.2016.03.002
Abstrakt: Residual host cellular DNA (rcDNA) is one of the principal risk associated with continuous cell lines derived medicines such as viral vaccines. To assess rcDNA degradation, we suggest two quantitative real-time PCR assays designed to separately quantify target sequences shorter and longer than the 200 bp risk limit, the relative abundance of both targets reflecting the extent of rcDNA fragmentation. The conserved multicopy ribosomal 18S RNA gene was targeted to detect host cell templates from most mammalian cell substrates commonly used in the manufacture of human viral vaccines. The detection range of the method was assessed on purified DNA templates from different animal origins. The standard calibrator origin and structural conformation were shown crucial to achieve accurate quantification. Artificial mixtures of PCR products shorter and longer than 200 bp were used as a model to check the ability of the assay to estimate the fragment size distribution. The method was successfully applied to a panel of Vero cell derived vaccines and could be used as a universal method for determination of both content and size distribution of rcDNA in vaccines.
(Copyright © 2016 International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.)
Databáze: MEDLINE
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