Assaying prions in cell culture: the standard scrapie cell assay (SSCA) and the scrapie cell assay in end point format (SCEPA).

Assaying prions in cell culture: the standard scrapie cell assay (SSCA) and the scrapie cell assay in end point format (SCEPA).

ISBN:

1064-3745

Call Number:

18576147

Accession Number:

18576147

URL:

http://eutils.ncbi.nlm.nih.gov/entrez/eutils/elink.fcgi?dbfrom=pubmed&id=18576147&retmode=ref&cmd=prlinks

Keywords:

Adenosine Triphosphate; Animals; Biological Assay; Cell Count; Cells; Cultured; Freezing; Membranes; Artificial; Mice; Prions; Software; Time Factors; Trypan Blue;

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.