The prion organotypic slice culture assay--POSCA

Abstract

Methods enabling prion replication ex vivo are important for advancing prion science. However, few such technologies exist and many prion strains are intractable with them. Here, we describe a prion organotypic slice culture assay (POSCA), which allows for prion amplification and titration ex vivo under conditions that closely resemble intracerebral infection. Organotypic slices are incubated with infectious inoculum as free-floating sections, washed and cultured for up to 8 weeks. Slice cultures are a rich source of protein or RNA and allow for stringent comparisons between uninfected and prion-infected samples generated from the same mouse. Thirty-five days after contact with prions, cerebellar slices have amplified PrP(Sc) quantitatively similar to that seen in vivo, but accelerated fivefold. The POSCA detects replication of specific prion strains from disparate sources, including bovines and ovines, with variable efficiency. The culture procedure and prion infection can be performed in 8 h.