College Park, Maryland June 6 - 10 , 2004 |
M3-B1 (13:30): SANS Characterization of Standard Virus Particles for Clinical Diagnostic Assays (Invited)
S. Krueger (NIST Center for Neutron Research), I. Elashvili, C. Wick (Edgewood Chemical Biological Center, 5183 Blackhawk Road, Aberdeen Proving Ground, MD 21010-5424), C. O’Connell (Biotechnology Division, NIST, 100 Bureau Drive, Stop 8311, Gaithersburg, MD 20899-8311), D.A. Kuzmanovic (Biotechnology Division, NIST, 100 Bureau Drive, Stop 8311, Gaithersburg, MD 20899-8311; Geo-Centers, Inc, P.O. Box 68, Aberdeen Proving Ground, MD 21010-5242)
Small angle neutron scattering (SANS) has been used to characterize the structure of wild-type and genetically modified MS2 bacterial virus (phage) by examining its physical characteristics in solution. This research was initiated in response to scientific needs from the biomedical community. Technical advances in molecular biology have made the synthesis of proteins for basic or biomedical research routine. Although artificially produced, many of these proteins are enzymatically active. In the case of some virus proteins, they also retain the ability to self-assemble into complete virus-like particles (VLPs). The inability of modern molecular biological techniques alone to identify or physically characterize these VLPs has hampered their use in vaccine development and as standards in commercially available clinical diagnostic assays.
Presently, noninfectious, genetically modified forms of the MS2 phage are commercially available for use in basic research diagnostic assays for the HIV, Ebola, Borna, Hepatitis A, C, and G, Dengue, Enterovirus, West Nile, and Norwalk viruses, among others. However these standard virus particles are not commercially available for human biomedical diagnostic assays because their physical characteristics in solution have not been measured. In this work, SANS has been coupled with a novel virus counting instrument, the Integrated Virus Detection System (IVDS) to address the scientific analytical challenges associated with this new generation of biological standards and potential vaccine carrying agents. Specifically, IVDS and the SANS contrast variation technique were employed to determine the molecular weight of the individual components of the MS2 virion (protein shell and genomic RNA) and the spatial relationship of the genomic RNA to its protein shell. Results will be shown for wild-type MS2 particles, recombinant MS2 particles containing RNA of different sizes and from different sources, and for RNA-free reconstituted MS2 capsids.
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