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Updated: Mar 6, 2026
Protein Misfolding Cyclic Amplification of Prions
Published on: November 7, 2012
In vitro amplification of H-type atypical bovine spongiform encephalopathy by protein misfolding cyclic amplification
Matthew J O'Connor1, Keith Bishop2, Robert G Workman1
1a School of Veterinary Medicine and Science, The University of Nottingham, Sutton Bonington Campus , College Road, Sutton Bonington , Leicestershire , UK.
Abstract:
The in vitro amplification of prions by serial protein misfolding cyclic amplification has been shown to detect PrPSc to 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 PrPSc from 3 distinct experimental isolates of H-BSE, could detect PrPSc in as little as 1×10-12 g 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 PrPSc with distinct protease K-resistance and protease K-resistant PrPSc molecular weights.
Insights
A new prion detection method, serial protein misfolding cyclic amplification (sPMCA), effectively detects atypical H-type Bovine Spongiform Encephalopathy (H-BSE) in cattle brain material. This highly sensitive and specific assay offers a faster alternative to traditional bioassays for H-BSE surveillance.
Area of Science:
- Veterinary Neurology
- Prion Disease Research
- Biochemical Assays
Background:
- Bovine spongiform encephalopathy (BSE) is a zoonotic prion disease in cattle.
- BSE occurs in classical (C-BSE) and two atypical forms: L-BSE and H-BSE.
- Atypical BSE forms are typically found in older, asymptomatic cattle and may arise spontaneously.
Purpose of the Study:
- To develop a serial protein misfolding cyclic amplification (sPMCA) method for detecting H-type BSE (H-BSE).
- To assess the sensitivity and specificity of the developed sPMCA assay for H-BSE detection.
Main Methods:
- Development of a novel sPMCA assay tailored for H-BSE detection.
- Testing the assay's ability to detect PrPSc from three distinct experimental H-BSE isolates.
- Evaluating the assay's sensitivity down to 1x10^-12 g of brain material.
Main Results:
- The sPMCA assay successfully detected PrPSc from all three experimental H-BSE isolates.
- The assay demonstrated high specificity and sensitivity, detecting PrPSc in minute quantities (1x10^-12 g).
- H-BSE amplification products were distinguishable from L-BSE (no amplification) and C-BSE (distinct protease resistance profiles).
Conclusions:
- The developed sPMCA method is a sensitive and specific tool for detecting H-BSE.
- This assay provides a rapid alternative to rodent bioassays for H-BSE surveillance.
- The sPMCA assay can differentiate between H-BSE, L-BSE, and C-BSE based on amplification and PrPSc characteristics.
