Limits...
The N-terminal sequence of prion protein consists an epitope specific to the abnormal isoform of prion protein (PrP(Sc)).

Masujin K, Kaku-Ushiki Y, Miwa R, Okada H, Shimizu Y, Kasai K, Matsuura Y, Yokoyama T - PLoS ONE (2013)

Bottom Line: Epitope analysis revealed that mAbs 8D5 and 6A12 recognized the PrP subregions between amino acids 31-39 and 41-47, respectively.This indicates that a PrP(Sc)-specific epitope exists in the N-terminal region of PrP(Sc), and mAbs 6A12 and 8D5 are powerful tools with which to detect native and intact PrP(Sc).We found that the ratio of proteinase K (PK)-sensitive PrP(Sc) to PK-resistant PrP(Sc) was constant throughout the disease time course.

View Article: PubMed Central - PubMed

Affiliation: Prion Disease Research Center, National Institute of Animal Health, Tsukuba, Ibaraki, Japan.

ABSTRACT
The conformation of abnormal prion protein (PrP(Sc)) differs from that of cellular prion protein (PrP(C)), but the precise characteristics of PrP(Sc) remain to be elucidated. To clarify the properties of native PrP(Sc), we attempted to generate novel PrP(Sc)-specific monoclonal antibodies (mAbs) by immunizing PrP-deficient mice with intact PrP(Sc) purified from bovine spongiform encephalopathy (BSE)-affected mice. The generated mAbs 6A12 and 8D5 selectivity precipitated PrP(Sc) from the brains of prion-affected mice, sheep, and cattle, but did not precipitate PrP(C) from the brains of healthy animals. In histopathological analysis, mAbs 6A12 and 8D5 strongly reacted with prion-affected mouse brains but not with unaffected mouse brains without antigen retrieval. Epitope analysis revealed that mAbs 8D5 and 6A12 recognized the PrP subregions between amino acids 31-39 and 41-47, respectively. This indicates that a PrP(Sc)-specific epitope exists in the N-terminal region of PrP(Sc), and mAbs 6A12 and 8D5 are powerful tools with which to detect native and intact PrP(Sc). We found that the ratio of proteinase K (PK)-sensitive PrP(Sc) to PK-resistant PrP(Sc) was constant throughout the disease time course.

Show MeSH

Related in: MedlinePlus

Immunoreactivity of mAbs 6A12 and 8D5 with PrPSc from different prion strains and animal species.(A) The brain homogenates [0.05% (w/v)] from ME7- and Chandler-affected mice were used. BSE: brain homogenate from BSE-affected mice. (B) Cross-reactivity of mAbs 6A12 and 8D5 with PrPSc from BSE and scrapie. Brain homogenates from unaffected [1% (w/v)] and affected [0.5% (w/v)] individuals were used. Total PrP in the brain homogenate was detected by routine western blotting (WB). BSE: brain homogenate from C-BSE-affected cattle; Scrapie: brain homogenate from scrapie-affected sheep. Molecular weight markers are shown on the left (kDa). The concentration of all mAbs for the immunoprecipitation assay was 1 µg/mL. MAbs 6A12 and 8D5 cross-reacted with PrPSc from different prion strains and animal species.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3585212&req=5

pone-0058013-g002: Immunoreactivity of mAbs 6A12 and 8D5 with PrPSc from different prion strains and animal species.(A) The brain homogenates [0.05% (w/v)] from ME7- and Chandler-affected mice were used. BSE: brain homogenate from BSE-affected mice. (B) Cross-reactivity of mAbs 6A12 and 8D5 with PrPSc from BSE and scrapie. Brain homogenates from unaffected [1% (w/v)] and affected [0.5% (w/v)] individuals were used. Total PrP in the brain homogenate was detected by routine western blotting (WB). BSE: brain homogenate from C-BSE-affected cattle; Scrapie: brain homogenate from scrapie-affected sheep. Molecular weight markers are shown on the left (kDa). The concentration of all mAbs for the immunoprecipitation assay was 1 µg/mL. MAbs 6A12 and 8D5 cross-reacted with PrPSc from different prion strains and animal species.

Mentions: The cross-reactivity of the newly generated mAbs to PrPSc from other animal species was analyzed. The mAbs 6A12 and 8D5 also detected PrPSc in the brains of ME7- and Chandler-affected mice (Fig. 2A), as well as PrPSc in the brains of scrapie-affected hamsters (data not shown), C-BSE-affected cattle, and scrapie-affected sheep (Fig. 2B). No PrP signal from healthy animals was detected (Figs. 2A, B).


The N-terminal sequence of prion protein consists an epitope specific to the abnormal isoform of prion protein (PrP(Sc)).

Masujin K, Kaku-Ushiki Y, Miwa R, Okada H, Shimizu Y, Kasai K, Matsuura Y, Yokoyama T - PLoS ONE (2013)

Immunoreactivity of mAbs 6A12 and 8D5 with PrPSc from different prion strains and animal species.(A) The brain homogenates [0.05% (w/v)] from ME7- and Chandler-affected mice were used. BSE: brain homogenate from BSE-affected mice. (B) Cross-reactivity of mAbs 6A12 and 8D5 with PrPSc from BSE and scrapie. Brain homogenates from unaffected [1% (w/v)] and affected [0.5% (w/v)] individuals were used. Total PrP in the brain homogenate was detected by routine western blotting (WB). BSE: brain homogenate from C-BSE-affected cattle; Scrapie: brain homogenate from scrapie-affected sheep. Molecular weight markers are shown on the left (kDa). The concentration of all mAbs for the immunoprecipitation assay was 1 µg/mL. MAbs 6A12 and 8D5 cross-reacted with PrPSc from different prion strains and animal species.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585212&req=5

pone-0058013-g002: Immunoreactivity of mAbs 6A12 and 8D5 with PrPSc from different prion strains and animal species.(A) The brain homogenates [0.05% (w/v)] from ME7- and Chandler-affected mice were used. BSE: brain homogenate from BSE-affected mice. (B) Cross-reactivity of mAbs 6A12 and 8D5 with PrPSc from BSE and scrapie. Brain homogenates from unaffected [1% (w/v)] and affected [0.5% (w/v)] individuals were used. Total PrP in the brain homogenate was detected by routine western blotting (WB). BSE: brain homogenate from C-BSE-affected cattle; Scrapie: brain homogenate from scrapie-affected sheep. Molecular weight markers are shown on the left (kDa). The concentration of all mAbs for the immunoprecipitation assay was 1 µg/mL. MAbs 6A12 and 8D5 cross-reacted with PrPSc from different prion strains and animal species.
Mentions: The cross-reactivity of the newly generated mAbs to PrPSc from other animal species was analyzed. The mAbs 6A12 and 8D5 also detected PrPSc in the brains of ME7- and Chandler-affected mice (Fig. 2A), as well as PrPSc in the brains of scrapie-affected hamsters (data not shown), C-BSE-affected cattle, and scrapie-affected sheep (Fig. 2B). No PrP signal from healthy animals was detected (Figs. 2A, B).

Bottom Line: Epitope analysis revealed that mAbs 8D5 and 6A12 recognized the PrP subregions between amino acids 31-39 and 41-47, respectively.This indicates that a PrP(Sc)-specific epitope exists in the N-terminal region of PrP(Sc), and mAbs 6A12 and 8D5 are powerful tools with which to detect native and intact PrP(Sc).We found that the ratio of proteinase K (PK)-sensitive PrP(Sc) to PK-resistant PrP(Sc) was constant throughout the disease time course.

View Article: PubMed Central - PubMed

Affiliation: Prion Disease Research Center, National Institute of Animal Health, Tsukuba, Ibaraki, Japan.

ABSTRACT
The conformation of abnormal prion protein (PrP(Sc)) differs from that of cellular prion protein (PrP(C)), but the precise characteristics of PrP(Sc) remain to be elucidated. To clarify the properties of native PrP(Sc), we attempted to generate novel PrP(Sc)-specific monoclonal antibodies (mAbs) by immunizing PrP-deficient mice with intact PrP(Sc) purified from bovine spongiform encephalopathy (BSE)-affected mice. The generated mAbs 6A12 and 8D5 selectivity precipitated PrP(Sc) from the brains of prion-affected mice, sheep, and cattle, but did not precipitate PrP(C) from the brains of healthy animals. In histopathological analysis, mAbs 6A12 and 8D5 strongly reacted with prion-affected mouse brains but not with unaffected mouse brains without antigen retrieval. Epitope analysis revealed that mAbs 8D5 and 6A12 recognized the PrP subregions between amino acids 31-39 and 41-47, respectively. This indicates that a PrP(Sc)-specific epitope exists in the N-terminal region of PrP(Sc), and mAbs 6A12 and 8D5 are powerful tools with which to detect native and intact PrP(Sc). We found that the ratio of proteinase K (PK)-sensitive PrP(Sc) to PK-resistant PrP(Sc) was constant throughout the disease time course.

Show MeSH
Related in: MedlinePlus