| A highly sensitive immunoassay for the detection of prion-infected material in whole human blood without the use of proteinase K. | |
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MedLine Citation:
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PMID: 20561299 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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BACKGROUND: The causal association of variant Creutzfeldt-Jakob disease (vCJD) with bovine spongiform encephalopathy has raised significant concerns for public health. Assays for vCJD infection are vital for the application of therapeutics, for the screening of organ donations, and to maintain a safe blood supply. Currently the best diagnostic tools for vCJD depend upon the detection of disease-associated prion protein (PrP(Sc) ), which is distinguished from normal background PrP (PrP(C) ) by proteinase K (PK) digestion, which can also degrade up to 90% of the target antigen. STUDY DESIGN AND METHODS: A sandwich enzyme-linked immunosorbent assay method was developed using unique antibodies for the detection of disease-associated PrP in the absence of PK treatment. In combination with immunoprecipitation the assay was optimized for the detection of pathogenic PrP in large volumes of whole blood. RESULTS: Optimization of the assay allowed detection of 2×10(4) LD(50) units/mL spiked in whole blood. Application of the assay to clinically relevant volumes enabled the detection of 750 LD(50) units/mL in 8mL of whole blood. CONCLUSION: By combining the use of a unique antibody that selectively immunoprecipitates PrP(Sc) with glycoform-restrictive antibodies we have developed a rapid assay for vCJD infection that does not require any PK treatment to achieve high levels of specificity in whole human blood, the most challenging potential analyte. The sensitivity of detection of vCJD infection is greater than the equivalent of a more than 2.5 million-fold dilution of infected brain, providing a highly sensitive immunoassay compatible with blood screening. |
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Authors:
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M Howard Tattum; Samantha Jones; Suvankar Pal; Azedeh Khalili-Shirazi; John Collinge; Graham S Jackson |
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Publication Detail:
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Type: Evaluation Studies; Journal Article; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: Transfusion Volume: 50 ISSN: 1537-2995 ISO Abbreviation: Transfusion Publication Date: 2010 Dec |
Date Detail:
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Created Date: 2010-12-03 Completed Date: 2011-01-20 Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 0417360 Medline TA: Transfusion Country: United States |
Other Details:
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Languages: eng Pagination: 2619-27 Citation Subset: IM |
Copyright Information:
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© 2010 American Association of Blood Banks. |
Affiliation:
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MRC Prion Unit and the Department of Neurodegenerative Disease, Institute of Neurology, University College London, London, UK. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Animals Antibodies, Monoclonal / diagnostic use, immunology, metabolism Antibody Specificity / drug effects Blood Safety Brain / pathology Donor Selection / methods, standards Endopeptidase K / metabolism*, pharmacology Enzyme-Linked Immunosorbent Assay / methods, standards Humans Immunoassay / methods, standards Mass Screening / methods, standards Mice Prion Diseases / blood, diagnosis*, pathology Prions / analysis, isolation & purification, metabolism Protein Isoforms / analysis, immunology Sensitivity and Specificity Serologic Tests / methods, standards |
| Grant Support | |
ID/Acronym/Agency:
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//Medical Research Council |
| Chemical | |
Reg. No./Substance:
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0/Antibodies, Monoclonal; 0/Prions; 0/Protein Isoforms; EC 3.4.21.64/Endopeptidase K |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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