{"id":739,"date":"2026-04-01T10:02:28","date_gmt":"2026-04-01T10:02:28","guid":{"rendered":"http:\/\/www.rischool.org\/?p=739"},"modified":"2026-04-01T10:02:28","modified_gmt":"2026-04-01T10:02:28","slug":"after-clonal-cells-were-established-they-were-transferred-to-new-plates-and-tested-for-permissiveness-to-infection-using-the-ssca","status":"publish","type":"post","link":"https:\/\/www.rischool.org\/?p=739","title":{"rendered":"\ufeffAfter clonal cells were established, they were transferred to new plates and tested for permissiveness to infection using the SSCA"},"content":{"rendered":"

\ufeffAfter clonal cells were established, they were transferred to new plates and tested for permissiveness to infection using the SSCA. == Western blotting. can be detectedex vivo. Despite the Rov9 and MovS6 cell lines being of different biological origin, they were both permissive and resistant to contamination with the same isolates of natural sheep scrapie as detected by SSCA. Rov9 subclones that are 20 occasions more sensitive than Rov9 to SSBP\/1-like scrapie contamination were isolated, but all the subclones maintained their resistance to isolates that failed to transmit to the parental line. The most sensitive subclone of the Rov9 cell line was used to estimate the infectious titer of a scrapie brain Ppia<\/a> pool (RBP1) and proved to be more sensitive than the mouse bioassay using wild-type mice. Increasing the sensitivity of the Rov9 ML264 cell line to SSBP\/1 contamination did not correlate with broadening susceptibility, as the specificity of permissiveness and resistance to other scrapie isolates was maintained. Prion diseases are a group of neurodegenerative diseases affecting humans and animals, including scrapie in sheep and goats and bovine spongiform encephalopathy (BSE) in cattle. A feature of prion diseases and, in particular, of scrapie, is the presence of different strains (6) which influence pathology and is most probably related to the conformation of the pathogenic form of the prion protein (PrPSc). The susceptibility of sheep to scrapie is determined by the PrP genotype; codons 136, 154, and 171 determine relative resistance and susceptibility, with amino acids valine (V), arginine (R), and glutamine (Q) at these positions (known as VRQ) being considered the sheep PrP allele most susceptible to classical scrapie (3). An array of diagnostic assessments exist for prion diseases, aimed at the detection of the disease-associated protease-resistant ML264<\/a> form of the naturally occurring PrPCprotein, termed PrPScor PrPresafter partial protease digestion. However, the level of detectable PrPScdoes not quantitatively correlate with prion infectivity (2) and the current biochemical analysis of PrPSccannot usually determine the strain (6,7). Mouse bioassay remains the gold standard for determining proof of infectivity, strain type, and infectious titer estimate in ruminant transmissible spongiform encephalopathy (TSE) research. Conventional mouse bioassays using wild-type mice are generally slow (>150 days, and considerably longer, >600, days for obtaining infectious titer information) and require multiple mice to be dosed (typically 6 or more) at each dilution of infectious material. Therefore, the development of an approach usingex vivocell-based assays ML264 remains an ethically and economically desirable option. Using cell lines permissive to mouse-passaged scrapie strains, Klhn et al. have developed a cell-based assay for measuringde novoinfection and the titer of mouse-passaged scrapie (18). The main limitation of adopting a cell-based approach is the scarcity of cell lines permissive to contamination with ML264 natural TSE strains (for a review, see recommendations31and34), as the majority of permissive cell lines can only be infected with rodent-adapted strains of scrapie and BSE (4,9,16,20,23,24,29,33,36). While there are currently no cell lines reported to be permissive to bovine BSE or human TSE diseases, there are cell lines which express ovine PrP that have been shown to be permissive to natural scrapie contamination (1,35). There is also one fibroblast-like deer cell line that is able to propagate chronic wasting disease (27). Two of the sheep scrapie-susceptible cell lines are the MovS6 cell line (1), a Schwann cell line derived from the tg301 transgenic mouse, and the Rov9 cell line (35), based on a stably transfected rabbit kidney epithelial cell line (RK13) that does not express endogenous PrP. Both express the VRQ allele of ovine PrP, the latter upon induction with doxycycline (35). These cell lines were found to be permissive to contamination with a PrP genotype-matched VRQ homozygous scrapie field case, andde novoPrPScmaintained its phenotype when used as an inoculum in mouse bioassays (1,35). Using fluorescence-activated cell sorting, Falanga et al. isolated Rov9 subclones that produce higher levels of PrPCand PrPScthan the parental cell.<\/p>\n","protected":false},"excerpt":{"rendered":"

\ufeffAfter clonal cells were established, they were transferred to new plates and tested for permissiveness to infection using the SSCA….<\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"closed","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[10],"tags":[],"class_list":["post-739","post","type-post","status-publish","format-standard","hentry","category-opioid"],"_links":{"self":[{"href":"https:\/\/www.rischool.org\/index.php?rest_route=\/wp\/v2\/posts\/739","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/www.rischool.org\/index.php?rest_route=\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/www.rischool.org\/index.php?rest_route=\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/www.rischool.org\/index.php?rest_route=\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/www.rischool.org\/index.php?rest_route=%2Fwp%2Fv2%2Fcomments&post=739"}],"version-history":[{"count":1,"href":"https:\/\/www.rischool.org\/index.php?rest_route=\/wp\/v2\/posts\/739\/revisions"}],"predecessor-version":[{"id":740,"href":"https:\/\/www.rischool.org\/index.php?rest_route=\/wp\/v2\/posts\/739\/revisions\/740"}],"wp:attachment":[{"href":"https:\/\/www.rischool.org\/index.php?rest_route=%2Fwp%2Fv2%2Fmedia&parent=739"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/www.rischool.org\/index.php?rest_route=%2Fwp%2Fv2%2Fcategories&post=739"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/www.rischool.org\/index.php?rest_route=%2Fwp%2Fv2%2Ftags&post=739"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}