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Rapid typing of transmissible spongiform encephalopathy strains with differential ELISA.

Simon S, Nugier J, Morel N, Boutal H, Créminon C, Benestad SL, Andréoletti O, Lantier F, Bilheude JM, Feyssaguet M, Biacabe AG, Baron T, Grassi J - Emerging Infect. Dis. (2008)

Bottom Line: Based on the particular biochemical properties of the abnormal prion protein (PrPsc) associated with BSE, and particularly the increased degradation induced by proteinase K in the N terminal part of PrPsc, we have developed a rapid ELISA designed to distinguish BSE from other scrapie strains.This assay clearly discriminates experimental ovine BSE from other scrapie strains and was used to screen 260 transmissible spongiform encephalopathy (TSE)-infected small ruminant samples identified by the French active surveillance network (2002/2003).In this context, this test has helped to identify the first case of natural BSE in a goat and can be used to classify TSE isolates based on the proteinase K sensitivity of PrPsc.

View Article: PubMed Central - PubMed

Affiliation: Commissariat à l'Energie Atomique, Gif-sur-Yvette, France.

ABSTRACT
The bovine spongiform encephalopathy (BSE) agent has been transmitted to humans, leading to variant Creutzfeldt-Jakob disease. Sheep and goats can be experimentally infected by BSE and have been potentially exposed to natural BSE; however, whether BSE can be transmitted to small ruminants is not known. Based on the particular biochemical properties of the abnormal prion protein (PrPsc) associated with BSE, and particularly the increased degradation induced by proteinase K in the N terminal part of PrPsc, we have developed a rapid ELISA designed to distinguish BSE from other scrapie strains. This assay clearly discriminates experimental ovine BSE from other scrapie strains and was used to screen 260 transmissible spongiform encephalopathy (TSE)-infected small ruminant samples identified by the French active surveillance network (2002/2003). In this context, this test has helped to identify the first case of natural BSE in a goat and can be used to classify TSE isolates based on the proteinase K sensitivity of PrPsc.

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Proteinase K (PK) sensitivity of Nor98 isolates in stringent and mild detergent conditions. The ELISA typing test was performed on Nor98 isolates, with 5 concentrations (0.4–1.1 µg per mg of tissue) in the stringent A′reagent (A) or in the mild A′′ reagent, adapted for PK-sensitive strains (B) (see Experimental Procedures). A/A′(or A/A′′) ratios were calculated for each PK concentration, and normalized by dividing by the A/A′ratio (or A/A′′) obtained for the experimental ovine bovine spongiform encephalopathy (BSE) sample at the maximal PK concentration. In the A’ reagent, even at the lowest PK concentration (PK 0.4 μg/mg tissue), the normalized ratios (using the experimental ovine BSE A/A′PK1.1 ratio) obtained for the Nor98 isolates are >1, thus being 3× more sensitive than experimental ovine BSE. To evaluate possible differences in PK sensitivity among Nor98 isolates, this experiment was reproduced with the A′′ reagent (panel B), which is 3- to 6-fold more protective than the A′reagent, as shown by the corresponding normalized ratios (A′or A′′ reagent) for the same PK concentration (1.1 μg PK/mg of tissue).
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Figure 6: Proteinase K (PK) sensitivity of Nor98 isolates in stringent and mild detergent conditions. The ELISA typing test was performed on Nor98 isolates, with 5 concentrations (0.4–1.1 µg per mg of tissue) in the stringent A′reagent (A) or in the mild A′′ reagent, adapted for PK-sensitive strains (B) (see Experimental Procedures). A/A′(or A/A′′) ratios were calculated for each PK concentration, and normalized by dividing by the A/A′ratio (or A/A′′) obtained for the experimental ovine bovine spongiform encephalopathy (BSE) sample at the maximal PK concentration. In the A’ reagent, even at the lowest PK concentration (PK 0.4 μg/mg tissue), the normalized ratios (using the experimental ovine BSE A/A′PK1.1 ratio) obtained for the Nor98 isolates are >1, thus being 3× more sensitive than experimental ovine BSE. To evaluate possible differences in PK sensitivity among Nor98 isolates, this experiment was reproduced with the A′′ reagent (panel B), which is 3- to 6-fold more protective than the A′reagent, as shown by the corresponding normalized ratios (A′or A′′ reagent) for the same PK concentration (1.1 μg PK/mg of tissue).

Mentions: After adapting the conditions of the PK treatment in the second set of measurements (A′conditions), we observed (see legend, Figure 6) a much lower A/A′ratio for those Nor-98, which enables discrimination of highly sensitive PK samples (nos. 24 and 26, Appendix Figure 2 and Table 3) to mildly sensitive PK samples (nos. 8, 11, 16, and 22).


Rapid typing of transmissible spongiform encephalopathy strains with differential ELISA.

Simon S, Nugier J, Morel N, Boutal H, Créminon C, Benestad SL, Andréoletti O, Lantier F, Bilheude JM, Feyssaguet M, Biacabe AG, Baron T, Grassi J - Emerging Infect. Dis. (2008)

Proteinase K (PK) sensitivity of Nor98 isolates in stringent and mild detergent conditions. The ELISA typing test was performed on Nor98 isolates, with 5 concentrations (0.4–1.1 µg per mg of tissue) in the stringent A′reagent (A) or in the mild A′′ reagent, adapted for PK-sensitive strains (B) (see Experimental Procedures). A/A′(or A/A′′) ratios were calculated for each PK concentration, and normalized by dividing by the A/A′ratio (or A/A′′) obtained for the experimental ovine bovine spongiform encephalopathy (BSE) sample at the maximal PK concentration. In the A’ reagent, even at the lowest PK concentration (PK 0.4 μg/mg tissue), the normalized ratios (using the experimental ovine BSE A/A′PK1.1 ratio) obtained for the Nor98 isolates are >1, thus being 3× more sensitive than experimental ovine BSE. To evaluate possible differences in PK sensitivity among Nor98 isolates, this experiment was reproduced with the A′′ reagent (panel B), which is 3- to 6-fold more protective than the A′reagent, as shown by the corresponding normalized ratios (A′or A′′ reagent) for the same PK concentration (1.1 μg PK/mg of tissue).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2570920&req=5

Figure 6: Proteinase K (PK) sensitivity of Nor98 isolates in stringent and mild detergent conditions. The ELISA typing test was performed on Nor98 isolates, with 5 concentrations (0.4–1.1 µg per mg of tissue) in the stringent A′reagent (A) or in the mild A′′ reagent, adapted for PK-sensitive strains (B) (see Experimental Procedures). A/A′(or A/A′′) ratios were calculated for each PK concentration, and normalized by dividing by the A/A′ratio (or A/A′′) obtained for the experimental ovine bovine spongiform encephalopathy (BSE) sample at the maximal PK concentration. In the A’ reagent, even at the lowest PK concentration (PK 0.4 μg/mg tissue), the normalized ratios (using the experimental ovine BSE A/A′PK1.1 ratio) obtained for the Nor98 isolates are >1, thus being 3× more sensitive than experimental ovine BSE. To evaluate possible differences in PK sensitivity among Nor98 isolates, this experiment was reproduced with the A′′ reagent (panel B), which is 3- to 6-fold more protective than the A′reagent, as shown by the corresponding normalized ratios (A′or A′′ reagent) for the same PK concentration (1.1 μg PK/mg of tissue).
Mentions: After adapting the conditions of the PK treatment in the second set of measurements (A′conditions), we observed (see legend, Figure 6) a much lower A/A′ratio for those Nor-98, which enables discrimination of highly sensitive PK samples (nos. 24 and 26, Appendix Figure 2 and Table 3) to mildly sensitive PK samples (nos. 8, 11, 16, and 22).

Bottom Line: Based on the particular biochemical properties of the abnormal prion protein (PrPsc) associated with BSE, and particularly the increased degradation induced by proteinase K in the N terminal part of PrPsc, we have developed a rapid ELISA designed to distinguish BSE from other scrapie strains.This assay clearly discriminates experimental ovine BSE from other scrapie strains and was used to screen 260 transmissible spongiform encephalopathy (TSE)-infected small ruminant samples identified by the French active surveillance network (2002/2003).In this context, this test has helped to identify the first case of natural BSE in a goat and can be used to classify TSE isolates based on the proteinase K sensitivity of PrPsc.

View Article: PubMed Central - PubMed

Affiliation: Commissariat à l'Energie Atomique, Gif-sur-Yvette, France.

ABSTRACT
The bovine spongiform encephalopathy (BSE) agent has been transmitted to humans, leading to variant Creutzfeldt-Jakob disease. Sheep and goats can be experimentally infected by BSE and have been potentially exposed to natural BSE; however, whether BSE can be transmitted to small ruminants is not known. Based on the particular biochemical properties of the abnormal prion protein (PrPsc) associated with BSE, and particularly the increased degradation induced by proteinase K in the N terminal part of PrPsc, we have developed a rapid ELISA designed to distinguish BSE from other scrapie strains. This assay clearly discriminates experimental ovine BSE from other scrapie strains and was used to screen 260 transmissible spongiform encephalopathy (TSE)-infected small ruminant samples identified by the French active surveillance network (2002/2003). In this context, this test has helped to identify the first case of natural BSE in a goat and can be used to classify TSE isolates based on the proteinase K sensitivity of PrPsc.

Show MeSH
Related in: MedlinePlus