Document Detail


An improved plaque reduction virus neutralization assay for human metapneumovirus.
MedLine Citation:
PMID:  17420056     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Virus-neutralizing antibodies against human metapneumovirus (hMPV) have been shown to be important indicators for protection in experimental animal models. An improved plaque reduction virus neutralization assay to detect hMPV-specific neutralizing antibodies was designed using two prototype recombinant hMPV strains expressing green fluorescent protein (GFP). These prototypes represented each of the main antigenic variants of hMPV, because antigenic variability could have implications for vaccine development. The utility of mutations in the F gene resulting in trypsin-independent replication was also tested. Although these mutant hMPV strains could replicate in the absence of trypsin, bigger plaque size was achieved with the addition of trypsin. Insertion of the GFP gene in the genome of hMPV did not affect replication of the virus in vitro. Plaques could be detected by measuring expression of GFP after 5 days by automated scanning. Ferret, hamster, and macaque sera positive for hMPV were compared in a conventional virus neutralization assay and the plaque reduction virus neutralization assay. The results obtained with the two assays were in agreement but the improved plaque reduction virus neutralization assay was faster, more suitable for high throughput testing, and 10-fold more sensitive than the conventional virus neutralization assay.
Authors:
Miranda de Graaf; Sander Herfst; Eefje J A Schrauwen; Bernadette G van den Hoogen; Albert D M E Osterhaus; Ron A M Fouchier
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Publication Detail:
Type:  Comparative Study; Evaluation Studies; Journal Article     Date:  2007-04-08
Journal Detail:
Title:  Journal of virological methods     Volume:  143     ISSN:  0166-0934     ISO Abbreviation:  J. Virol. Methods     Publication Date:  2007 Aug 
Date Detail:
Created Date:  2007-07-02     Completed Date:  2007-08-28     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  8005839     Medline TA:  J Virol Methods     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  169-74     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Animals
Antibodies, Viral / blood*
Cercopithecus aethiops
Cricetinae
Ferrets
Genes, Reporter
Green Fluorescent Proteins / analysis,  genetics
Macaca
Metapneumovirus / genetics,  immunology*,  physiology
Neutralization Tests / methods*
Plaque Assay
Recombination, Genetic
Sensitivity and Specificity
Serum / immunology
Trypsin / metabolism
Vero Cells
Viral Proteins / genetics
Virus Replication / genetics,  physiology
Chemical
Reg. No./Substance:
0/Antibodies, Viral; 0/Viral Proteins; 147336-22-9/Green Fluorescent Proteins; EC 3.4.21.4/Trypsin

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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