Dean migration mechanisms: facilitating the control of microbial contaminants in cell therapy product manufacturing

Cell therapy products are rapidly transforming clinical practice, but their short shelf-lives and inability to undergo terminal sterilization create major challenges for sterility testing. Conventional rapid microbiological methods (RMMs) are hindered by the dense cellular background of therapeutic samples, which masks rare microbial contaminants and necessitates pre-analytical processing. Efficient separation of microorganisms from high-density cell suspensions is therefore a critical prerequisite for enabling real-time, in-process sterility assurance. Here, we systematically elucidate the Dean flow-dominated migration mechanism and determine its effective range for continuous, label-free separation of non-typical contaminants ≤ 5 μm in microchannels exceeding 40 μm in height. We demonstrate that particles with ap/h < 0.05 undergo exclusive Dean-induced lateral migration, while those near the inertial focusing threshold (ap/h ≈ 0.07) exhibit a Reynolds number-dependent transition between unfocused and centerline-focused streams. Leveraging these principles, we designed optimized channel geometries that achieved > 95% separation efficiency and > 96% purity of T cells versus three morphologically distinct bacteria at 10⁵ bacteria/mL. At ultra-low loads (< 10 CFU/mL), culture-based assays confirmed 100% detection for inocula > 1 CFU/mL. Our findings validate Dean migration as a governing mechanism for submicron particle separation and provide a path toward integrating microfluidic modules into closed CAR-T manufacturing platforms, advancing real-time microbial quality control in cell therapy production.