Abstract
Human induced pluripotent stem cells (hiPSCs) must be manufactured as advanced therapy medicinal products (ATMPs) for innovative tissue replacement clinical applications. Yet, production of hiPSCs under current Good Manufacturing Practice (cGMP) presents many hurdles, such as the large-scale cell expansion needed to reach therapeutically-relevant hiPSC doses. For the monitoring of this phase, a fast and reliable cell counting method should be used. Conventional manual cell counting by the hemocytometer method is dependent on the operator's expertise and is time-consuming. Therefore, automation of sample preparation and analysis is needed to improve precision and rapidity of hiPSC cell counting. We investigated whether an automated cell counting method could be validated for use with hiPSCs, in comparison with a reference cell counting method included in the European Pharmacopeia, 10th edition. The proposed method was the fluorescence imaging-based NucleoCounter NC-100 system, whereas the reference method was manual cell counting using a Bürker hemocytometer. The validation strategy complied with EudraLex cGMP regulations for ATMP manufacturing and ICH Q2(R1) indications for validation of analytical methods. The use of the NucleoCounter NC-100 system for automated cell counting was validated, focusing on accuracy, specificity, intra- and inter-operator reproducibility, range and linearity, showing higher precision than the manual method. The automated method can be used more effectively than the manual one for hiPSC cell counting. Thus, this piece of work paves the way for all cGMP facilities that want to pursue hiPSC manufacturing for clinical use.