Serum, Methods and Considerations for Use

Most transection reagents recommend transfecting in serum reduced or serum free medium. There are some that are advertised to be equally as effective in the presence or absence of serum. I would be inclined to transfect serum-free to be on the safe side. Unfortunately the only way to prove anything is through testing. If this was going to become a model for transfection that would be done routinely, it might be worth investing the time. If the transfection is specific to a short-term project, it may not be worth the effort - just go serum free to reduce the number of variables. The cell line you are working with is relatively robust and should be able to take a period of serum free transfection fairly well.

To my knowledge, there is no animal serum product approved by an entity such as the FDA.Sera products from the larger suppliers are generally manufactured in facilities compliant with GMPs and ISO standards. Manufacturers may also conform to other international quality standards. Each manufacturer generally with have product lines offering a variety of testing levels. A review of the specification and/or C of A prior to any purchase is a key activity to ensure the product is suitable for the intended application. Definitely work with the manufacturer to obtain the information needed to make the right purchase. They can inform you of product label claims, compliance to quality standards, testing and origin of the products.

There are many functions that FBS supplies. Which ones are most important likely relies on the application. It provides attachment factors, growth factors, simple nutrients like amino acids and vitamins, stability factors for nutrients through binding, etc.The list is a very long one. These are the reasons it is difficult stop using it. I makes the culture of cells relatively simple and stable through providinges so many functions in one material.There is still more to learn about its functions in cell culture.

Newborn calf serum (or calf serum) can be utilized in cell culture for a few different reasons. If a process is very price sensitive, it makes sense to design using a less expensive product. The newborn calf serum is generally more stable in price and availability compared to FBS if reliability is a key concern. Newborn calf serum is useful in applications where the cell line is robust and cell attachment is the primary goal. Newborn calf serum may not be a good choice where development resources are limited. The culture system will be a little more challenging to design. I do not think I would attempt to substitute newborn calf serum in an existing process utilizing FBS unless there are adequate resources for development. There is a substantial amount of work to be done to verify the system has not been significantly altered by the change.

Time spent in qualification of assays for any application can pay off substantially in the long run. Designing assays for qualification of sera have to be based in the specific application. There may be more than one assay required to manage the risk for a process. Characteristics may include parameters such as cell attachment, population doubling time, viral infectivity, etc. It may be worth running through some sort of fault analysis, such as a process FMEA, to make sure all the critical points are identified. Design several assays and see which has the best assessment qualities for the specific application. Ideally it would be one short assay, but that is not always achievable. The variability in cell culture processes can make it challenging to create a single assay for multiple applications. Building up the statistical confidence in the assay is also a key function in minimizing risk. A potential pitfall is in creating assays that are very sensitive. It really needs to fit the application. If 90% of the samples pass the test and are all effective in the application – that is great. There is no value in being more discriminating when it is not necessary.

Finding precipitates in serum is not uncommon. They can be composed of proteins, lipids, salts and any combination of these.

It should not affect the performance of the serum at all. Precipitates can be minimized by careful thawing of the serum. Do not overexpose to heat and mix gently while thawing. Keep the number of freeze thaw cycles to a minimum.

Heat inactivation of sera was originally implemented to inactivate complement which interfered in assays. The process has impacts on other molecules that may not have been quantified or evaluated. The literature varies as to the potential positive and negative effects of heat inactivation. For most cell lines in routine culture, it is likely not needed. The question becomes less clear in specific applications such as proteomic studies. The only way to ensure heat inactivation is not required is data from controlled experiments. It may be determined that heat inactivation is required for consistency with prior data, culture performance or not at all. If it is determined that heat inactivation is required, I would advise that heat inactivated serum be purchased. Manufacturers should have validated processes for heat inactivation that will ensure the product you receive is as consistent as possible.

In reference specifically to MSC culture, there are products qualified for MSC applications that are not heat inactivated on the market. There are some culture routines and literature which support using heat inactivated serum in MSC culture. The final decision on to use heat inactivated serum or not is specific to the application.