Limitations of Manual Cultivation
High variability and low consistency are major problems in manual stem cell culture processes, for both growth of stem cells and their differentiation into pre-defined cell types. Variations in environmental conditions during bioprocessing have the potential to significantly impact the outcome of stem cell cultures, making it difficult to reproduce experimental protocols and interpret the results obtained.
Many manual handling steps are needed to obtain the final cell population, requiring multiple passages, media exchanges, and cell transfers, each with the potential for contamination and propagation of errors.
Most significantly, it is practically impossible to control several key parameters during these manipulations—most notably temperature and CO2 concentration—as plates are moved from environmentally controlled incubators at 37ºC and 5% CO2, to a biological safety cabinet at room temperature (20–24ºC) and atmospheric CO2 levels (~0.04%), as illustrated in Figure 1.
As a consequence, rapid cooling and diffusion of CO2 out of the media occur, leading to fluctuations in the pH of the culture (Figure 2). Stem cells are extremely sensitive to such changes in pH, and this leads to poor reproducibility in manual culturing, representing a major obstacle to both furthering our understanding of stem cell biology and the commercialization of stem cell applications.