Name: protein misfolding cyclic amplification of prions
Uploaded: 2012-11-07T12:00:00
Description: Watch this Scientific Journal Video about Protein Misfolding Cyclic Amplification of Prions at JoVE.com

우리는 원고의 중요한 읽기에 대한 박사 개밥 바라기 철 마리 라모스 감사드립니다. 이 작품은 연구 자원에 대한 국립 센터 (P20 RR0115635-6, C06 RR17417-01 및 G20RR024001)과 국립 신경 질환 연구소와 스트로크 (2R01 NS052609)에 의해 지원되었다.

1. 장비 준비 <ol><li> Misonix 3000이나 37 일정 온도를 유지하기 위해 열 전자 Neslab EX-7 물 목욕 (이케아, NH)에 연결 Misonix 4000 sonicator (Farmingdale, NY) ° C.를 사용하여 열 과학 (Waltham, MA)에서 얻은 돔 캡 200 μl 얇은 벽 PCR 관 스트립의 샘플을 Sonicate. </li><li> 새로운 sonicator는 지속적인 작동 9 &#39;브레이크에서 &quot;기간이 필요합니다. 두 달간 브레이크의 기간은 10로 설정 진폭 수준 40 초 sonication 버?...

<ol>
	<li>Ayers, J. I., Schutt, C. R., Shikiya, R. A., Aguzzi, A., Kincaid, A. E., Bartz, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+strain-encoded+relationship+between+PrP+replication,+stability+and+processing+in+neurons+is+predictive+of+the+incubation+period+of+disease.">The strain-encoded relationship between PrP replication, stability and processing in neurons is predictive of the incubation period of disease.</a> PLoS pathogens. 7, e1001317 (2011).</li><li>Barria, M. A., Mukherjee, A., Gonzalez-Romero, D., Morales, R., Soto, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=De+novo+generation+of+infectious+prions+in+vitro+produces+a+new+disease+phenotype.">De novo generation of infectious prions in vitro produces a new disease phenotype.</a> PLoS Pathog. 5, e1000421 (2009).</li><li>Bessen, R. A., Marsh, R. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biochemical+and+physical+properties+of+the+prion+protein+from+two+strains+of+the+transmissible+mink+encephalopathy+agent.">Biochemical and physical properties of the prion protein from two strains of the transmissible mink encephalopathy agent.</a> J. Virol. 66, 2096-2101 (1992).</li><li>Bessen, R. A., Marsh, R. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Distinct+PrP+properties+suggest+the+molecular+basis+of+strain+variation+in+transmissible+mink+encephalopathy.">Distinct PrP properties suggest the molecular basis of strain variation in transmissible mink encephalopathy.</a> J. Virol. 68, 7859-7868 (1994).</li><li>Bessen, R. A., Marsh, R. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+of+two+biologically+distinct+strains+of+transmissible+mink+encephalopathy+in+hamsters.">Identification of two biologically distinct strains of transmissible mink encephalopathy in hamsters.</a> J. Gen. Virol. 73, 329-334 (1992).</li><li>Castilla, J., Gonzalez-Romero, D., Saa, P., Morales, R., De Castro, J., Soto, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Crossing+the+species+barrier+by+PrP(Sc)+replication+in+vitro+generates+unique+infectious+prions.">Crossing the species barrier by PrP(Sc) replication in vitro generates unique infectious prions.</a> Cell. 134, 757-768 (2008).</li><li>Castilla, J., Morales, R., Saa, P., Barria, M., Gambetti, P., Soto, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell-free+propagation+of+prion+strains.">Cell-free propagation of prion strains.</a> EMBO J. 27, 2557-2566 (2008).</li><li>Caughey, B., Raymond, G. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+scrapie-associated+form+of+PrP+is+made+from+a+cell+surface+precursor+that+is+both+protease-+and+phospholipase-sensitive.">The scrapie-associated form of PrP is made from a cell surface precursor that is both protease- and phospholipase-sensitive.</a> J. Biol. Chem. 266, 18217-18223 (1991).</li><li>Deleault, N. R., Harris, B. T., Rees, J. R., Supattapone, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Formation+of+native+prions+from+minimal+components+in+vitro.">Formation of native prions from minimal components in vitro.</a> Proceedings of the National Academy of Sciences of the United States of America. 104, 9741-9746 (2007).</li><li>Dickinson, A. G., Fraser, H., Meikle, V. M., Outram, G. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Competition+between+different+scrapie+agents+in+mice.">Competition between different scrapie agents in mice.</a> Nat. New Biol. 237, 244-245 (1972).</li><li>Gonzalez-Romero, D., Barria, M. A., Leon, P., Morales, R., Soto, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Detection+of+infectious+prions+in+urine.">Detection of infectious prions in urine.</a> FEBS Lett. 582, 3161-3166 (2008).</li><li>Green, K. M., Castilla, J., Seward, T. S., Napier, D. L., Jewell, J. E., Soto, C., Telling, G. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Accelerated+high+fidelity+prion+amplification+within+and+across+prion+species+barriers.">Accelerated high fidelity prion amplification within and across prion species barriers.</a> PLoS Pathog. 4, e1000139 (2008).</li><li>Hadlow, W. J., Race, R. E., Kennedy, R. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temporal+distribution+of+transmissible+mink+encephalopathy+virus+in+mink+inoculated+subcutaneously.">Temporal distribution of transmissible mink encephalopathy virus in mink inoculated subcutaneously.</a> J. Virol. 61, 3235-3240 (1987).</li><li>Kocisko, D. A., Come, J. H., Priola, S. A., Chesebro, B., Raymond, G. J., Lansbury, P. T., Caughey, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell-free+formation+of+protease-resistant+prion+protein.">Cell-free formation of protease-resistant prion protein.</a> Nature. 370, 471-474 (1994).</li><li>Kurt, T. D., Telling, G. C., Zabel, M. D., Hoover, E. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trans-species+amplification+of+PrP(CWD)+and+correlation+with+rigid+loop+170N.">Trans-species amplification of PrP(CWD) and correlation with rigid loop 170N.</a> Virology. 387, 3235-3240 (2009).</li><li>Maddison, B. C., Baker, C. A., Terry, L. A., Bellworthy, S. J., Thorne, L., Rees, H. C., Gough, K. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Environmental+sources+of+scrapie+prions.">Environmental sources of scrapie prions.</a> J. Virol. 84, 11560-11562 (2010).</li><li>Nichols, T. A., Pulford, B., Wyckoff, A. C., Meyerett, C., Michel, B., Gertig, K., Hoover, E. A., Jewell, J. E., Telling, G. C., Zabel, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Detection+of+protease-resistant+cervid+prion+protein+in+water+from+a+CWD-endemic+area.">Detection of protease-resistant cervid prion protein in water from a CWD-endemic area.</a> Prion. 3, 171-183 (2009).</li><li>Prusiner, S. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+proteinaceous+infectious+particles+cause+scrapie.">Novel proteinaceous infectious particles cause scrapie.</a> Science. 216, 136-144 (1982).</li><li>Saborio, G. P., Permanne, B., Soto, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sensitive+detection+of+pathological+prion+protein+by+cyclic+amplification+of+protein+misfolding.">Sensitive detection of pathological prion protein by cyclic amplification of protein misfolding.</a> Nature. 411, 810-813 (2001).</li><li>Saunders, S. E., Bartz, J. C., Vercauteren, K. C., Bartelt-Hunt, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+enzymatic+treatment+of+soil-bound+prions+effectively+inhibits+replication.">An enzymatic treatment of soil-bound prions effectively inhibits replication.</a> Appl. Environ. Microbiol. 77, 4313-4317 (2011).</li><li>Saunders, S. E., Shikiya, R. A., Langenfeld, K., Bartelt-Hunt, S. L., Bartz, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Replication+efficiency+of+soil-bound+prions+varies+with+soil+type.">Replication efficiency of soil-bound prions varies with soil type.</a> Journal of virology. , (2011).</li><li>Schutt, C. R., Bartz, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prion+interference+with+multiple+prion+isolates.">Prion interference with multiple prion isolates.</a> Prion. 2, 61-63 (2008).</li><li>Shikiya, R. A., Ayers, J. I., Schutt, C. R., Kincaid, A. E., Bartz, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Co-infecting+prion+strains+compete+for+a+limiting+cellular+resource.">Co-infecting prion strains compete for a limiting cellular resource.</a> Journal of. 84, 5706-5714 (2010).</li><li>Shikiya, R. A., Bartz, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+generation+of+high+titer+prions.">In vitro generation of high titer prions.</a> Journal of virology. , (2011).</li><li>Weber, P., Giese, A., Piening, N., Mitteregger, G., Thomzig, A., Beekes, M., Kretzschmar, H. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generation+of+genuine+prion+infectivity+by+serial+PMCA.">Generation of genuine prion infectivity by serial PMCA.</a> Veterinary microbiology. 123, 346-357 (2007).</li></ol>

증폭기 전염성 prion 단백질의 과제는 생체 실험의 긴 잠복 기간 및 비용입니다. PMCA 기술은 전염성 prion 요원을 증폭 할 수있는 비용 효과적인 수단입니다. 여러 실험실 정확하게 체외 7, 9, 12, 14, 19,24에 prion의 긴장을 증폭 할 수 PMCA의 능력을 확인하고 있습니다. Prion 질환은 종 사이에 전염 될 수 있습니다. Bessen 및 습지 효과적으로 두 개의 햄스터 ?...

<table border="1"><tbody><tr><td> 시약 / 장비 </td><td> 제조업 자 </td><td> 고양이. 수 </td></tr><tr><td> Misonix 3000 </td><td> Misonix </td><td> S-3000 </td></tr><tr><td> Misonix 4000 </td><td> Misonix </td><td> S-4000 </td></tr><tr><td> Tenbroeck의 조직 그라인더 </td><td> Kontes </td><td> 885000-0007 </td></tr><tr><td> Neslab EX-7 물 목욕 </td><td> 열 전자 </td><td> Neslab EX-7 </td></tr><tr><td> 0.2 ML PCR 튜브 스트립 </td><td> 열 과학 </td><td> AB-0451 </td></tr><tr><td> 트리톤 X-100 </td><td> 시그마 알드리치 </td><td> T9284-100ML </td></tr><tr><td> 전체 프로테아제 억제제 </td><td> 로슈 </td><td> 11 697 498 001 </td></tr><tr><td> EDTA (에틸렌 다이아 민 테트라 초산) </td><td> JT 베이커 </td><td> 4040-00 </td></tr><tr><td> DPBS </td><td> Mallinckrodt 베이커 Mediatech </td><td> 21-031 CV </td></tr><tr><td> Versi - 드라이 랩 Soakers </td><td> 피셔 과학 </td><td> 14 206 28 </td></tr><tr><td> Repti 칼로리 히터 </td><td> 동물원 메드 랩 주식회사 </td><td> RH-4 </td></tr></tbody></table>

protein misfolding cyclic amplification of prions

Prions 동물과 인간 9,18에 필연적으로 치명적인 전도 할 수있는 해면상 뇌병증 (TSE)을 일으킬 전염성 대리인입니다. prion 단백질은 두 개의 isoforms, 비 전염성 호스트 인코딩 단백질 (PRP C) 및 전염성 단백질 (PRP SC), PRP C 8 비정상적으로 - 접힌 이소 형이 있습니다. prion 에이전트 작업의 과제 중 하나는 이전의 호스트 접종 13 다음 임상 증상의 발전에 긴 잠복기입니다. 이 전통적으로 길고 비싼 동물 bioassay 연구를 위임. 또한, PRP SC의 생화학 및 biophysical 속성이 제대로 자신의 특이한 형태와 집계 상태로 인해 특징을 가지고 있습니다. PRP SC는 체외 14 PRP SC에 PRP C의 씨앗 변환 할 수 있습니다. PMCA은 adva 소요의 체외 기술이다PRP C와 PRP SC의 씨앗 19 분 금액을 초과하는 금액을 포함하는 시스템에서, 가속 속도로, PRP SC 많은 양의 생산 sonication과 부화주기를 사용하여이 능력을 ntage. 이 기술은 효과적으로 prion 변형 간섭을 에뮬레이션하기 위해 PRP C에서 PRP SC 전환의 종과 변형 특성을 요점을 되풀이하고, 감염된 조직, 체액, 환경 샘플 6,7,16에서 PRP SC의 매우 낮은 수준을 증폭하기 위해 증명되었습니다 23. , 오염을 최소화 일관된 결과를 생성하고, 그 결과를 정량화에 대한 권장 사항을 포함 본 논문은 내용 PMCA 프로토콜을. 우리는 또한 전염성 prion의 변종, prion 변형 간섭 및 환경에서 prions의 감지의 생성 및 특성 등 여러 PMCA 응용 프로그램을 논의합니다.

단백질 misfolding의 순환 증폭 (PMCA)는 체외 7 PRP SC, 12, 14, 19, 24을 증폭하는 데 사용됩니다. 성공적으로 PRP SC 증폭은 그림 3과 같이 PK 방지 prion 단백질 (햄스터 파생 prion의 변종이 19 ~ 30 kDa를 마이그레이션)의 서양 blots에 밴드 강도의 증가에 의해 표시됩니다.의 밴드 강도의 증가는 이후 PMCA는 PK 방지 PRP SC 소재의 증폭을 나타냅니다. ?...

Watch this Scientific Journal Video about Protein Misfolding Cyclic Amplification of Prions at JoVE.com

Protein Misfolding Cyclic Amplification of Prions

단백질 misfolding의 순환 증폭 (PMCA)는 prion 변환과 변형 및 종 장벽의 연구에서 시험 관내 분석이다. 또한 prion 검출 분석으로 사용할 수 있습니다.

Prions의 단백질 Misfolding의 순환 증폭

Prions  are infectious agents that cause the inevitably fatal transmissible spongiform  encephalopathy (TSE) in animals and humans9,18. The prion protein ...

Research

JoVE Journal

Immunology and Infection

A Comparative Approach to Characterize the Landscape of Host-Pathogen Protein-Protein Interactions

호스트 병원체 단백질 - 단백질 상호 작용의 풍경의 특성을 비교 접근

Significant efforts were gathered to generate  large-scale comprehensive protein-protein interaction network maps. This is  instrumental to understand the ...

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Infectious prion (PrPRes) material is likely the cause of fatal, neurodegenerative transmissible spongiform encephalopathy (TSE) diseases1. Transmission ...

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The RT-QuIC technique is a sensitive in vitro cell-free prion amplification assay based mainly on the seeded misfolding and aggregation of recombinant ...

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Targeted therapies against the human epidermal growth factor receptor 2 (HER2) have radically changed the outcome of patients with HER2-positive breast ...

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The use of recombinant viruses has become crucial in basic or applied virology. Reverse genetics has been proven to be an extremely powerful technology, ...