Tips for selecting the “right” ELISPOT filter plate and making the best protocol choices

 Nitrocellulose or PVDF? There are two different types of membranes (in 96-Well plate formats) that are used in ELISPOT applications: Nitrocellulose (NC) and polyvinylidene fluoride (PVDF). The reason that these are the predominant membrane types used in ELISPOT is based largely on their fortuitous suitability in non-ELISPOT applications rather than on the development of optimized assay substrates to address the specific needs of the ELISPOT assay. NC and then PVDF membranes were developed to serve the needs of sterile filtration applications. As was later discovered, the porosity and binding properties of these two membrane types (NC and PVDF) enabled them to be used in two extremely important research applications; DNA and protein blotting. Finally, in response to the specific requirements of molecular biology and protein chemistry applications, NC and PVDF membrane-bottomed 96-well plates were developed and made commercially available. Independently, separately, and at about the same time, ELISPOT assays were developed on 96-well plastic plates and took advantage of enzyme-linked immunosorbent assay (ELISA) techniques that had been perfected in that format. Since fundamentally the immunodetection component of ELISPOT assays and protein (i.e., Western) blotting is essentially identical, it was only a matter of time until the overwhelming majority of ELISPOT assays were performed on membrane-bottomed, 96-well plates.