Suarez: One of the common missteps that I have encountered when working with customers is a lack of mindfulness of the sensitivity or fragility of cells. This can manifest in results that are inconsistent or unexpected.
An important starting place is to ensure you have a robust and validated cell bank, and that cell viability has been checked. Tracking the passage number can also help minimize variability. Throughout the cell expansion phase, it is important to be aware of cell stressors. Ensure that the freezing and thawing process is quality controlled, and that variability is assessed throughout. Do not grow your cells too densely before passaging, as this can put unnecessary stress on cells, leading to inconsistent or unexpected results.
In cryopreservation, some of the missteps we have seen include lab personnel using double Styrofoam packaging or repurposing Styrofoam packaging materials in efforts to control a freeze-thaw cycle. Now, we have CoolCell® products available that are much more controlled to help ensure you have a quality freeze process to produce a quality and robust master cell bank.
Siler: Mindfulness is key. I tell customers frequently to be mindful of the protein concentration of their extracellular matrix, such as Matrigel®. This is a biological product, and the protein concentration will differ from lot to lot. Sometimes customers assume that every lot has the exact same concentration. I encourage them to pay attention to the lot number. By being mindful of the protein concentration for a particular lot number, you can ensure greater consistency from experiment to experiment and ensure that you are always using the same concentration.
Mogen: One area I have seen difficulty in is the handling of scale-up vessels, such as multi-layer vessels like Corning® CellSTACK® and HYPERStack® technology. These are polystyrene plastic vessels that are often used in scaling up cell culture and bioproduction.
They are large, and when filled with media they can be heavy, so people often do not realize their fragility. I have run into some examples of our clients handling the vessels in an indelicate way, leading to breakages. We train our clients to be careful with those vessels by making sure that they are unpackaging them and laying them down carefully on the lab bench or in the biosafety cabinet, as they can crack and break. We also do extensive validation around the packaging.
Suarez: When we engage with customers, often the question of whether cell culture vessels can be re-used will be raised. If the vessel is made of single-use plastic, then they are single-use by design, and there are reasons for that. The consistency features rely on factors such as surface treatment, and if you attempt to reuse them, those surfaces can be altered or diminished. Cells excrete extracellular matrix on the surface as they grow, thus changing the surface. Continued use is going to introduce process variability. Morphological changes seen in cells can often be indicative of molecular changes that impact overall quality and productivity.
An attempt to reuse a single-use vessel can indicate that a customer has strayed from the best practice standard. We want to minimize variability so that you have consistent results, and your experiments are reproducible and robust. Overall, it is going to save you time and cost that you may otherwise invest in trying to optimize that variability.