High density bioprocessing of human pluripotent stem cells by metabolic control and in silico modeling.

To harness the full potential of human pluripotent stem cells (hPSCs) we combined instrumented stirred tank bioreactor (STBR) technology with the power of in silico process modeling to overcome substantial, hPSC-specific hurdles toward their mass production. Perfused suspension culture (3D) of matrix-free hPSC aggregates in STBRs was applied to identify and control process-limiting parameters including pH, dissolved oxygen, glucose and lactate levels, and the obviation of osmolality peaks provoked by high density culture. Media supplements promoted single cell-based process inoculation and hydrodynamic aggregate size control.

Modeling methodology for defining a priori the hydrodynamics of a dynamic suspension bioreactor. Application to human induced pluripotent stem cell culture.

Three-dimensional dynamic suspension is becoming an effective cell culture method for a wide range of bioprocesses, with an increasing number of bioreactors proposed for this purpose. The complex hydrodynamics establishing within these devices affects bioprocess outcomes and efficiency, and usually expensive in vitro trial-and-error experiments are needed to properly set the working parameters. Here we propose a methodology to define a priori the hydrodynamic working parameters of a dynamic suspension bioreactor, selected as a test case because of the complex hydrodynamics characterizing its operating condition.

Sensitivity of human pluripotent stem cells to insulin precipitation induced by peristaltic pump-based medium circulation: considerations on process development.

Controlled large-scale production of human pluripotent stem cells (hPSCs) is indispensable for their envisioned clinical translation. Aiming at advanced process development in suspension culture, the sensitivity of hPSC media to continuous peristaltic pump-based circulation, a well-established technology extensively used in hydraulically-driven bioreactors, was investigated. Unexpectedly, conditioning of low protein media (i.

Impact of Feeding Strategies on the Scalable Expansion of Human Pluripotent Stem Cells in Single-Use Stirred Tank Bioreactors.

Unlabelled: : The routine application of human pluripotent stem cells (hPSCs) and their derivatives in biomedicine and drug discovery will require the constant supply of high-quality cells by defined processes. Culturing hPSCs as cell-only aggregates in (three-dimensional [3D]) suspension has the potential to overcome numerous limitations of conventional surface-adherent (two-dimensional [2D]) cultivation. Utilizing single-use instrumented stirred-tank bioreactors, we showed that perfusion resulted in a more homogeneous culture environment and enabled superior cell densities of 2.

Cardiac differentiation of human pluripotent stem cells in scalable suspension culture.

Cardiomyocytes (CMs) generated from human pluripotent stem cells (hPSCs) are a potential cell source for regenerative therapies, drug discovery and disease modeling. All these applications require a routine supply of relatively large quantities of in vitro-generated CMs. This protocol describes a suspension culture-based strategy for the generation of hPSC-CMs as cell-only aggregates, which facilitates process development and scale-up.

Controlling expansion and cardiomyogenic differentiation of human pluripotent stem cells in scalable suspension culture.

To harness the potential of human pluripotent stem cells (hPSCs), an abundant supply of their progenies is required. Here, hPSC expansion as matrix-independent aggregates in suspension culture was combined with cardiomyogenic differentiation using chemical Wnt pathway modulators. A multiwell screen was scaled up to stirred Erlenmeyer flasks and subsequently to tank bioreactors, applying controlled feeding strategies (batch and cyclic perfusion).