ISO/PWI 26125-2

Biotechnology Tumorigenicity Testing — Part 2: Nucleic Acid Detection for Undifferentiated Pluripotent Stem Cells in cell and gene therapy products ISO/PWI 26125-2

Scope

This document specifies the general requirements for detecting residual pluripotent stem cells in cell therapy products using reverse transcription quantitative polymerase chain reaction (RT-qPCR) or reverse transcription digital polymerase chain reaction (RT-dPCR) assays, with messenger RNA (mRNA), long none cording RNA (lncRNA) or microRNA (miRNA) as target analytes. It provides guidance on:
 — Sample representativity, including the considerations in sample size；sample replicates；nucleic acid quality control;
— Assay selection and design , including selection of fit-for-purpose test methods; control setup; target gene selection, reference gene selection, normalization strategies, Primer, and Probe Design; and Amplification Strategy
— Method validation, covering general requirements, determination of the limit of detection (LOD), and revalidation;
— Data analysis and reporting, covering quality control for assay data; normalization of data; result interpretation and reporting.
This document is applicable to the development and validation of RT-qPCR or RT-dPCR as methods intended for the detection of residual pluripotent stem cells in cell therapy products.