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Despite considerable regulatory and clinical hurdles, the development and use of cell-based therapies are gaining momentum. As more of these therapies move toward commercial approval and larger-scale distribution, associated manufacturing and processing technologies are being advanced. Modern technologies directed at downstream processing seek to distribute such therapies from the manufacturing site to the patient more efficiently and reliably. Novel small-scale downstream solutions boost the transformation of cell therapies from abstraction to reality.

Cryopreservation is the process by which intact living cells are preserved at cryogenic temperatures in liquid nitrogen. Freezing cells allow them to be stored, often for years, in a state where their normal metabolic activity is suspended in order to protect them from damage due to chemical reactivity and time. The cells can then be thawed and resuscitated as needed. Cells are cryopreserved in order to guard against genetic drift in continuous cell lines, or against transformation or differentiation in noncontinuous cell lines, such as stem cells and primary cells. In the case of primary cells that are isolated directly from the tissue of interest, they have a finite ability to divide; therefore, cryopreservation is necessary to preserve their unique characteristics for future experiments. Optimal cryopreservation of cells relies on proper freezing and thawing methods. To be protected from structural damage during the freeze-thaw process, mammalian cells are frozen in the presence of cryoprotectant. Post-thaw viability assays are then conducted to measure the success of the cryopreservation techniques by calculating the percentage of frozen cells that are alive and able to recover normal function once thawed. Key Concepts: Cryopreservation is used to enable long-term storage, preserve current genetic state, prevent transformation or differentiation during subculture and provide a back-up stock in case of infection/contamination during culture. A controlled rate of freezing and rapid thawing is necessary for optimal cryopreservation and recovery. Care must be taken to minimize transient warming events during transfer and storage, as it impacts viability and recovery. Active thawing results in higher cell viability and recovery than passive thawing. Timing is critical to all stages of the cryopreservation process. All materials should be ready before beginning the procedure, and steps should be taken to ensure each sample is handled with minimal delay. Method standardisation and quality control are necessary to maintain sample integrity and reproducibility. Keywords: cryopreservation; controlled-rate freezing, cryoprotectant; viability; freezing; cell thawing

personnel and process flows, and resulting air flow patterns. Within the BSC critical zone, sites should be in close proximity to operations such as centrally located settle plates and particle sensors not more than 1 foot from the work site. Surface viable sites should be in close proximity to processing activity, at locations contacted by operator gloves, areas where materials transfer from lower classification, and other facility surfaces (e.g. floors, walls) to assess disinfectant efficacy. Suggested sampling frequencies are shown in Tables 1 and 2. EM Levels: Cell therapy areas typically will have ISO 5 BSCs placed within an ISO 7 room with ISO 8 support such as a gowning airlock. This classification scheme is outlined in USP Cellular and Tissue-Based Products. Suggested EM levels for cell therapy aseptic processing are shown in Table 3. Conclusion: Considerations are offered to help cell therapy manufacturers to set-up and an appropriate risk-based EM sampling program that meets criteria of being meaningful, manageable, and defendable.