ELISPOT Frequently Asked Questions (FAQ)



	Do I need to use sterile plates? Typical ELISPOT assays involve maximally 1-2 days of cell culture. Antibiotics in the culture medium typically suffice to avoid infections. Still, using sterile plates is a wise precaution.		Other FAQ sections: Assay Performance Scanning/ScAnalysis ELISPOT Analyzers Image Quality GLP compliance
	What can I do if my cell numbers are very low? For optimal results, T cells and antigen-presenting cells need to form a monolayer on the membrane. If T cells are limiting (e.g. when testing cells isolated form a solid organ or biopsy material) any number of T cells can be plated on a monolayer of APCs. If both cell types are limiting, CTL recommends using the low-volume plates manufactured by Whatman. Alternatively, the activation culture can be done in regular U- or V-bottom plates, followed by a transfer of the cells into the ELISPOT plate. (Pipet very gently to minimize shear forces during the transfer.) We can help customize your assay. Please contact CTL’s scientists through ctl-europe@immunospot.com.
	What type of membrane does CTL recommend? In general, PVDF plates are recommended. The fractal surface and the hydrophobic properties of PVDF membranes are ideal for the binding of most coating antibodies and substrates. To receive our plate type info flyer, please contact ctl-europe@immunospot.com.
	Can I also use clear plates in ImmunoSpot® analyzers? Yes, ImmunoSpot® analyzers can handle clear plates. However, in general, CTL recommends opaque plates as these do not conduct light, thereby avoiding light reflections that interfere with image analysis.
	At the moment, I use Agarose plates. Can these be analyzed as well? ImmunoSpot® S4 Analyzers are capable of analyzing Agarose overlay Elispots. However, Agarose spots tend to fade very quickly, and the light reflections on the Agarose make the analysis process more labor intensive.
	Can I use ImmunoSpot® systems to analyze clear bottom plates with "golden" spots (that is, spots developed using the gold-silver staining system)? The ImmunoSpot® analyzers can handle these plates without any problems.
	How long can I store packaged sterile ELISPOT plates? Uncoated, sterile packaged PVDF plates, if undamaged, have a shelf life of several years.
	How long can I store coated wet plates? Coated, wet plates are stable for about one week if kept at 4°C. They may be dried and stored longer, but the stability of different antibodies differs considerably.
	Can the developed plates be autoclaved and re-used? No. However, if only a part of a plate was used for an assay, the unused wells can be re-used. (Use antibiotics. They provide sufficient protection for sterility in a 1-2 day assay).