In vitro amplification of H-type atypical bovine spongiform encephalopathy by protein misfolding cyclic amplification.

Abstract

The in vitro amplification of prions by serial protein misfolding cyclic amplification has been shown to detect PrPto levels at least as sensitive as rodent bioassay but in a fraction of the time. Bovine spongiform encephalopathy is a zoonotic prion disease in cattle and has been shown to occur in 3 distinct forms, classical BSE (C-BSE) and 2 atypical BSE forms (L-BSE and H-BSE). Atypical forms are usually detected in asymptomatic, older cattle and are suggested to be spontaneous forms of the disease. Here, we show the development of a serial protein misfolding cyclic amplification method for the detection of H-BSE. The assay could detect PrPfrom 3 distinct experimental isolates of H-BSE, could detect PrPin as little as 1×10g of brain material and was highly specific. Additionally, the product of serial protein misfolding cyclic amplification at all dilutions of seed analyzed could be readily distinguished from L-BSE, which did not amplify, and C-BSE, which had PrPwith distinct protease K-resistance and protease K-resistant PrPmolecular weights.