Biologics Analytical R&D, Merck & Co., Inc. Kenilworth, New Jersey, USA.
J Pharm Biomed Anal. 2022 Mar 20;211:114564. doi: 10.1016/j.jpba.2021.114564. Epub 2022 Jan 6.
Biologics drug manufacturers need to show clearance of host cell DNA in the purified drug to alleviate safety concerns. Currently, sensitive methods of DNA quantification, like quantitative real-time PCR (qPCR) or digital PCR (dPCR) are used in different labs. We have developed an isothermal PCR method to detect Chinese Hamster Ovary (CHO) DNA and performed it in droplet digital PCR (ddPCR) format to make the method quantitative. In this novel approach the protein drug is directly added to the isothermal PCR reaction mix avoiding DNA extraction and related manipulations. Then nanoliter-sized droplets are generated, followed by incubation, and reading the fluorescence in each droplet. The method is simple and fast and can potentially be adapted to other dPCR systems.
生物制药商需要在纯化药物中清除宿主细胞 DNA，以减轻安全隐患。目前，不同实验室使用灵敏的 DNA 定量方法，如实时定量 PCR(qPCR)或数字 PCR(dPCR)。我们开发了一种等温 PCR 方法来检测中国仓鼠卵巢(CHO)DNA，并将其用于微滴式数字 PCR(ddPCR)格式，使该方法具有定量性。在这种新方法中，直接将蛋白药物添加到等温 PCR 反应混合物中，避免了 DNA 提取和相关操作。然后生成纳升级大小的微滴，接着孵育，并读取每个微滴中的荧光。该方法简单快速，并且可能适用于其他 dPCR 系统。
