Establishment of a protein misfolding cyclic amplification for PrPSc.
- Author:
Jun HAN
1
;
Lu HAN
;
Qi SHI
;
Song SHI
;
Xin WANG
;
Bao-Yun ZHANG
;
Xiao-Ping DONG
Author Information
- Publication Type:Journal Article
- MeSH: Animals; Biochemistry; methods; Blotting, Western; Brain; metabolism; pathology; Cricetinae; PrPSc Proteins; analysis; chemistry; Prion Diseases; diagnosis; metabolism; Protein Folding; Reproducibility of Results; Sensitivity and Specificity
- From: Chinese Journal of Experimental and Clinical Virology 2007;21(3):202-204
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo establish a methodology of protein misfolding cyclic amplification (PMCA) and utilize in the detection of PrP(Sc) in brain tissues from prion diseases.
METHODSDifferent amounts of Scrapie 263K agent bulk were mixed with brain homogenates of health hamsters and treated with repeated incubation/sonication for 10 to 15 cycles. The proteinase K-resistant PrP(Sc) was evaluated with Western Blot.
RESULTSIn this experimental situation, 263K agent replicated rapidly in vitro, utilizing hamsters' brains as the medium. With the established PrP(Sc)-PMCA technique, PrP(Sc) signals in the preparations containing less than 10(-5) diluted 263K bulk could be detected. Compared with conveniently used immuno-blot assay, the sensitivity of PrP(Sc)-PMCA for PrP(Sc) was 10(5) to 10(6)-fold increased. It has been also shown that homogenates of cerebellar and brain stem could be used as the medium for PrP(Sc) replication.
CONCLUSIONA rapidly replicating method for PrP(Sc), PrP(Sc)-PMCA, was successfully established, providing a new approach for early diagnosis of prion diseases and research on the biological features of prion.
