Scientific Background and Utility of Serum-free Media in ELISPOT.
Serum-free cell culture media allow users to standardize their cell culture conditions by avoiding the use of undefined and highly-variable serum products derived from humans or animals, e.g. human AB serum or fetal calf serum (FCS).
The high variability in the biological properties of different serum batches makes it necessary to pre-screen many batches in order to obtain a batch that is well-suited for a given application. Even a brief exposure of PBMC to a mitogenic serum batch, for example during washing or freezing of these cells, will result in a high background in cytokine assays. In addition, toxic or inhibitory serum batches will jeopardize the assay results. The unique performance of each serum batch necessitates the purchase of large lots to assure consistency of assay conditions as long as possible. This adds to the inconvenience and cost associated with the use of serum. Switching to or using a different batch of serum introduces a significant unpredictable variable to the test conditions each time, making test results difficult to compare. Furthermore, infectious risks associated with serum are leading to increasingly tighter restrictions for international shipments and exchange of any material that has been exposed to serum.
For all these reasons, there is considerable pressure from regulatory agencies and from the scientific community to avoid the use of serum and move to consistent, defined substitutes. However, because serum is naturally rich in a multitude of growth factors and other essentials for cell growth and functionality, it has been challenging to develop a serum substitute for primary cells in general, and for PBMC in particular. Most serum-free media contain enough additives to permit the growth of robust tumor cells, and hybridomas, but freshly-isolated human PBMC require more stringent conditions to maintain their viability and functionality in serum-free media.
CTL has been working for years with US governmental, industrial, and academic partners to standardize ELISPOT and other direct ex vivo cytokine assays performed on freshly-isolated or cryopreserved PBMC. These efforts have resulted in the CTL Serum-free Media Platform, which consists of three product lines that have been carefully designed to cover the four critical steps of working with human PBMC: washing, testing, freezing, and thawing. All four previously required the use of serum.