Western blot mapping of disease-specific amyloid in various animal species and humans with transmissible spongiform encephalopathies using a high-yield purification method

Beekes, Michael; Baldauf, Elizabeth; Cassens, Sven; Diringer, Heino; Keyes, P.; Scott, A. C.; Wells, G. A. H.; Brown, Paul; Gibbs, C. J.; Gajdusek, D. Carleton

doi:10.1099/0022-1317-76-10-2567

Cited by 58 publications

(38 citation statements)

References 71 publications

(43 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…SDS-PAGE and Western blot analyses were performed as described elsewhere (3,37) with recently published modifications (12). PrP signals were visualized on a X-OMAT AR (Kodak; Sigma-Aldrich, Steinheim, Germany) film.…”

Section: Western Blot Examinationsmentioning

confidence: 99%

“…Muscle and nerve specimens to be examined by Western blotting were processed as previously described (12) for the extraction of PrP Sc in the form of PrP27-30 (3,37); the final pellets were resuspended in 15 μl of 2× sample loading buffer and heated to 100°C for 5 minutes before Western blotting using the anti-PrP mAb 3F4 (38).…”

Section: Western Blot Examinationsmentioning

confidence: 99%

“…Proteinase K-digested homogenate from scrapie hamster brains, used as a PrP27-30 reference in the Western blotting analyses, was prepared as outlined previously (12,37).…”

Section: Western Blot Examinationsmentioning

confidence: 99%

See 2 more Smart Citations

Preclinical deposition of pathological prion protein PrPSc in muscles of hamsters orally exposed to scrapie

Thomzig¹,

Schulz‐Schaeffer²,

Kratzel³

et al. 2004

J. Clin. Invest.

View full text Add to dashboard Cite

Recently, pathological prion protein PrP Sc , the putative key constituent of infectious agents causing transmissible spongiform encephalopathies (TSEs), was found in muscles of rodents experimentally infected with scrapie and in patients with Creutzfeldt-Jakob disease (CJD). For the assessment of risk scenarios originating from these findings (e.g., alimentary transmission of pathogens associated with bovine spongiform encephalopathy [BSE]

show abstract

Section: Western Blot Examinationsmentioning

confidence: 99%

Section: Western Blot Examinationsmentioning

confidence: 99%

See 1 more Smart Citation

Preclinical deposition of pathological prion protein PrPSc in muscles of hamsters orally exposed to scrapie

Thomzig¹,

Schulz‐Schaeffer²,

Kratzel³

et al. 2004

J. Clin. Invest.

View full text Add to dashboard Cite

show abstract

“…All nerve samples were incubated at 37 • C for 60 min with 500 µL 0.25% collagenase in TBS buffer containing 2 mM CaCl 2 . After halting the reaction by adding 10 µL 250 mM EDTA, tissue-extraction of PrP Sc in the form of the protease resistant 27-30 kD fragment (PrP27-30) was initiated in line with a previously published protocol [6], including the use of proteinase K (PK) to destroy normal PrP C . The final pellets were resuspended with 20 µL of 2 × sample loading buffer and heated to 100 • C for 10 min prior to Western blotting using the monoclonal antiPrP antibody (mAb) 3F4 [27] and an alkaline phosphatase-conjugated secondary goat-anti-mouse antibody.…”

Section: Western Blotmentioning

confidence: 99%

“…The final pellets were resuspended with 20 µL of 2 × sample loading buffer and heated to 100 • C for 10 min prior to Western blotting using the monoclonal antiPrP antibody (mAb) 3F4 [27] and an alkaline phosphatase-conjugated secondary goat-anti-mouse antibody. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analyses were performed as described elsewhere [6,49]. Antibody binding was visualized using CDP-Star (Tropix) as a chemoluminescent substrate for alkaline phosphatase and a Kodak X-OMAT AR film, exposed for 30-60 min.…”

Section: Western Blotmentioning

confidence: 99%

Propagation of scrapie in peripheral nerves after footpad infection in normal and neurotoxin exposed hamsters

et al. 2007

View full text Add to dashboard Cite

-As is known from various animal models, the spread of agents causing transmissible spongiform encephalopathies (TSE) after peripheral infection affects peripheral nerves before reaching the central nervous system (CNS) and leading to a fatal end of the disease. The lack of therapeutic approaches for TSE is partially due to the limited amount of information available on the involvement of host biological compartments and processes in the propagation of the infectious agent. The in vivo model presented here can provide information on the spread of the scrapie agent via the peripheral nerves of hamsters under normal and altered axonal conditions. Syrian hamsters were unilaterally footpad (f.p.) infected with scrapie. The results of the spatiotemporal ultrasensitive immunoblot-detection of scrapie-associated prion protein (PrP Sc ) in serial nerve segments of both distal sciatic nerves could be interpreted as a centripetal and subsequent centrifugal neural spread of PrP Sc for this route of infection. In order to determine whether this propagation is dependent on main components in the axonal cytoskeleton (e.g. neurofilaments, also relevant for the component 'a' of slow axonal transport mechanisms), hamsters were treated -in an additional experiment-with the neurotoxin ß,ß'-iminodiproprionitrile (IDPN) around the beginning of the scrapie infection. A comparison of the Western blot signals of PrP Sc in the ipsilateral and in the subsequently affected contralateral sciatic nerve segments with the results revealed from IDPN-untreated animals at preclinical and clinical stages of the TSE disease, indicated similar amounts of PrP Sc . Furthermore, the mean survival time was unchanged in both groups. This in vivo model, therefore, suggests that the propagation of PrP Sc along peripheral nerves is not dependent on an intact neurofilament component of the axonal cytoskeleton. Additionally, the model indicates that the spread of PrP Sc is not mediated by the slow component 'a' of the axonal transport mechanism.scrapie / PrP Sc / nerve / footpad / IDPN

show abstract

Amyotrophy in Prion Diseases

Worrall¹,

Rowland²,

Chin³

et al. 2000

Arch Neurol

View full text Add to dashboard Cite

Amyotrophic lateral sclerosis was once thought to be caused by persistent viral infection, partly because some patients with transmissible Creutzfeldt-Jakob disease showed prominent amyotrophy. However, in the past 15 years there has been little interest in the amyotrophy in prion diseases, and the possible link to amyotrophic lateral sclerosis has been eschewed. We analyzed case reports of prion disease published after 1968 for evidence of amyotrophy. We defined amyotrophy as clinically evident fasciculation buttressed by electromyographic results in some cases. We sought evidence of motor neuron degeneration at autopsy. Prion disease was proved by transmissibility, immunohistochemistry demonstration of protease-resistant prion protein, or finding a mutation in the prion protein gene. Amyotrophy was noted in 27 patients: 13 with sporadic Creutzfeldt-Jakob disease, 2 with familial Creutzfeldt-Jakob disease, and 12 with Gerstmann-Sträussler-Scheinker disease. Of the 27, 23 showed clinical fasciculation and 10 had electromyographic evidence of denervation. The spinal cord was examined in 8 patients: 6 showed loss of motor neurons, 1 showed vacuolation of motor neurons, and 1 reported no abnormalities. Another 23 patients had typical histopathological characteristics but lacked molecular or biochemical proof of prion disease. The total number of patients with amyotrophy and proven prion disease that we identified was 50. This case review supports the belief that amyotrophy is occasionally a prominent feature of Creutzfeldt-Jakob disease and underscores the importance of documenting lower motor neuron function and the crucial role of examining the spinal cord at autopsy in cases of prion disease.

show abstract

Western blot mapping of disease-specific amyloid in various animal species and humans with transmissible spongiform encephalopathies using a high-yield purification method

Cited by 58 publications

References 71 publications

Preclinical deposition of pathological prion protein PrPSc in muscles of hamsters orally exposed to scrapie

Preclinical deposition of pathological prion protein PrPSc in muscles of hamsters orally exposed to scrapie

Propagation of scrapie in peripheral nerves after footpad infection in normal and neurotoxin exposed hamsters

Amyotrophy in Prion Diseases

Contact Info

Product

Resources

About