Is it time for a new standard for the culture of iPSCs?
For those delivering advanced therapy medicinal products, the question of assessing and managing cell population heterogeneity presents an ongoing challenge. Solentim and EverCell Bio addressed this topic at a recent webinar during which they discussed the use of induced pluripotent stem cells (iPSCs) and sharing data from EverCell Bio’s iPSC lines when moving into the clinic.
As cell line development processes are translated from the academic lab to a clinical setting, the need to change the process or adopt clinical-grade reagents can be daunting, and there are several key considerations that must be taken into account.
Implementing the right matrix is a fundamental step in successfully propagating iPSCs. Vitronectin and MatrigelTMare most frequently used but both have their shortcomings. One of the main bugbears applying to both is the need to pre-coat plates with these matrices, leading to extra time and loss of an expensive reagent if not all the wells are needed on a plate. Moreover, as Matrigel is a biologically-derived product, there is complexity in keeping uniformity across multiple lots. Vitronectin overcomes this by being a synthetic product derived from a well-defined recombinant protein.
Currently methodologies are further limited when it comes to proof of clonality. The classic method of cloning iPSC lines in dishes as closely packed cells in colonies creates problems in identifying the true clonality of a cell line. In turn, this may lead to low-level contamination by a cell line with an unwanted phenotype or differentiation propensity, leading to deviations away from the desired product for a therapeutic agent. For scientists wishing to image single cells to confirm that their cell lines are clonally derived, the current matrices provide challenges to capture a clear image of the single cell or require additional time to eyeball the single cell in the well. So, what is the solution?
A new standard
Solentim collaborated with EverCell Bio to demonstrate the benefits of using a liquid matrix, MatriCloneTM, recently released to the market, in combination with the VIPSTM, Solentim’s tried and tested single cell seeding device. The MatriClone VIPS combination was easy to implement across multiple iPSC lines and enabled EverCell Bio to cut its workflow timeline in half, leading to a banked clone in 20 days.
Factors such as pre-adapting the cells to MatriClone, starting cell concentration and the size of the VIPS droplets all were important to obtain a robust workflow.
Biological impact of a new reagent
The big question remains: ‘how will these cell culture reagents impact the pluripotency of an iPSC line and the later differentiation capacity?’ EverCell Bio demonstrated that the workflow was highly compatible with different cell culture media and cell lines. From the five iPSC lines analyzed, pluripotency markers remained intact and the karyotype remained normal after single-cell isolation in MatriClone using the VIPS.
When asked by an audience member whether differentiation was seen on the cloning plate, Philip Manos, Founder, President & Scientific Director of EverCell Bio, reported none was evident from the 60-plus plates processed in the study. After the expansion of a subset of clones via normal clustering growth, each cell line was exposed to the company’s tried and tested differentiation methodology for mixed cortical neurons where all the cell lines responded as expected and showed the expected markers. It was noted by Mr Manos that the quality of the input cell lines is key to obtain a strong and positive outcome. All in all, the Solentim process met or exceeded expectations at EverCell Bio.
This new standard is an exciting development for all scientists looking to be sure that their iPSC lines are derived from a single clone. Tune in to the webinar to find out more about Solentim’s robust workflow solution for cloning iPSCs and its potential to impact standards, consistency, and confidence of clonality between different laboratories in both academic and pharma research.
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