title: Assaying prions in cell culture: the standard scrapie cell assay (SSCA) and the scrapie cell assay in end point format (SCEPA).
abstract: Prions are usually quantified by bioassays based on intracerebral inoculation of animals, which are slow, imprecise, and costly. We have developed a cell-based prion assay that is based on the isolation of cell lines highly susceptible to certain strains (Rocky Mountain Laboratory and 22L) of mouse prions and a method for identifying individual, prion-infected cells and quantifying them. In the standard scrapie cell assay (SSCA), susceptible cells are exposed to prion-containing samples for 4 days, grown to confluence, passaged two or three times, and the proportion of rPrP(Sc)-containing cells is determined with automated counting equipment. The dose response is dynamic over 2 logs of prion concentrations. The SSCA has a standard error of +/-20-30%, is as sensitive as the mouse bioassay, 10 times faster, at least 2 orders of magnitude less expensive, and it is suitable for robotization. Assays performed in a more time-consuming end point titration format extend the sensitivity and show that infectivity titers measured in tissue culture and in the mouse are similar.
authors.affiliation: Department of Infectology, Scripps Florida, Jupiter, FL, USA.
authors.forename: Sukhvir P
authors.initials: SP
authors.lastname: Mahal
authors.forename: Cheryl A
authors.initials: CA
authors.lastname: Demczyk
authors.forename: Emery W
authors.initials: EW
authors.lastname: Smith
authors.forename: Peter-Christian
authors.initials: PC
authors.lastname: Klohn
authors.forename: Charles
authors.initials: C
authors.lastname: Weissmann
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001681:Biological Assay / Q000379:methods*; D002452:Cell Count; D002478:Cells, Cultured; D005615:Freezing; D008567:Membranes, Artificial; D051379:Mice; D011328:Prions / Q000032:analysis*; D012984:Software; D013997:Time Factors; D014343:Trypan Blue
chemical_list: D008567:Membranes, Artificial; D011328:Prions; D000255:Adenosine Triphosphate; D014343:Trypan Blue