Simple, specific, rapid, and pharmacopoeia-compliant qPCR approach for the detection of mycoplasma in biopharmaceuticals.

IF 4.7 2区医学 Q2 MEDICINE, RESEARCH & EXPERIMENTAL

Molecular Therapy-Methods & Clinical Development Pub Date : 2025-08-20 eCollection Date: 2025-09-11 DOI:10.1016/j.omtm.2025.101572

Sergio Dos Santos, Emilie Lespinasse, Baptiste Bonnet, Stéphane Basmaciogullari

{"title":"Simple, specific, rapid, and pharmacopoeia-compliant qPCR approach for the detection of mycoplasma in biopharmaceuticals.","authors":"Sergio Dos Santos, Emilie Lespinasse, Baptiste Bonnet, Stéphane Basmaciogullari","doi":"10.1016/j.omtm.2025.101572","DOIUrl":null,"url":null,"abstract":"<p><p>Cell substrate utilized in the production of biologics intended for human use need to be cleared from mycoplasma contaminant as described in pharmacopoeia. While the gold-standard method for mycoplasma detection involves lengthy microbiology and cell-based assays, nucleic acid technologies offer the possibility to accelerate testing. In this study, we develop a qPCR assay capable of specifically detecting 11 mycoplasma species relevant to pharmacopoeias with only two primers and two hydrolysis probes, which greatly facilitates operations. While the primers cross-react with bacterial species, the specificity conferred by the hydrolysis probes allows for confident interpretation of the qPCR result and unambiguous statement regarding the presence of mycoplasma in the test article. Amplicon sequencing can further confirm the identity of contaminants. This comprehensive assay can therefore be of great help to quality control laboratories embedded in biologics production sites, which need to provide results in a timely manner and contribute to root cause analysis in case of contamination.</p>","PeriodicalId":54333,"journal":{"name":"Molecular Therapy-Methods & Clinical Development","volume":"33 3","pages":"101572"},"PeriodicalIF":4.7000,"publicationDate":"2025-08-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12441703/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Molecular Therapy-Methods & Clinical Development","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1016/j.omtm.2025.101572","RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2025/9/11 0:00:00","PubModel":"eCollection","JCR":"Q2","JCRName":"MEDICINE, RESEARCH & EXPERIMENTAL","Score":null,"Total":0}

引用次数: 0

Abstract

Cell substrate utilized in the production of biologics intended for human use need to be cleared from mycoplasma contaminant as described in pharmacopoeia. While the gold-standard method for mycoplasma detection involves lengthy microbiology and cell-based assays, nucleic acid technologies offer the possibility to accelerate testing. In this study, we develop a qPCR assay capable of specifically detecting 11 mycoplasma species relevant to pharmacopoeias with only two primers and two hydrolysis probes, which greatly facilitates operations. While the primers cross-react with bacterial species, the specificity conferred by the hydrolysis probes allows for confident interpretation of the qPCR result and unambiguous statement regarding the presence of mycoplasma in the test article. Amplicon sequencing can further confirm the identity of contaminants. This comprehensive assay can therefore be of great help to quality control laboratories embedded in biologics production sites, which need to provide results in a timely manner and contribute to root cause analysis in case of contamination.

查看原文本刊更多论文

简便、特异、快速、符合药典要求的qPCR方法用于生物制药中支原体的检测。

如药典所述，用于人用生物制剂生产的细胞底物需要清除支原体污染物。虽然支原体检测的金标准方法涉及冗长的微生物学和基于细胞的测定，但核酸技术提供了加速检测的可能性。在本研究中，我们建立了一种qPCR方法，仅用两个引物和两个水解探针就能特异性检测11种与药典相关的支原体，大大方便了操作。当引物与细菌物种交叉反应时，水解探针赋予的特异性允许对qPCR结果进行自信的解释，并明确说明测试品中支原体的存在。扩增子测序可以进一步确认污染物的身份。因此，这种综合分析方法对生物制剂生产现场的质量控制实验室有很大帮助，这些实验室需要及时提供结果，并有助于在发生污染的情况下进行根本原因分析。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

求助全文

约1分钟内获得全文求助全文

来源期刊

Molecular Therapy-Methods & Clinical Development Biochemistry, Genetics and Molecular Biology-Molecular Biology

CiteScore

9.90

自引率

4.30%

发文量

163

审稿时长

12 weeks

期刊介绍： The aim of Molecular Therapy—Methods & Clinical Development is to build upon the success of Molecular Therapy in publishing important peer-reviewed methods and procedures, as well as translational advances in the broad array of fields under the molecular therapy umbrella. Topics of particular interest within the journal''s scope include: Gene vector engineering and production, Methods for targeted genome editing and engineering, Methods and technology development for cell reprogramming and directed differentiation of pluripotent cells, Methods for gene and cell vector delivery, Development of biomaterials and nanoparticles for applications in gene and cell therapy and regenerative medicine, Analysis of gene and cell vector biodistribution and tracking, Pharmacology/toxicology studies of new and next-generation vectors, Methods for cell isolation, engineering, culture, expansion, and transplantation, Cell processing, storage, and banking for therapeutic application, Preclinical and QC/QA assay development, Translational and clinical scale-up and Good Manufacturing procedures and process development, Clinical protocol development, Computational and bioinformatic methods for analysis, modeling, or visualization of biological data, Negotiating the regulatory approval process and obtaining such approval for clinical trials.