Immunochemical studies of tobacco mosaic virus—II

Regenmortel, M.H.V. Van; Hardie, Gwendoline

doi:10.1016/0019-2791(76)90326-8

Cited by 41 publications

(5 citation statements)

References 40 publications

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“…The theory predicts formation of virion oligomers in the region of antigen excess, of large aggregates in balanced virus-antibody mixtures, and of single virions, surrounded by as many antibody molecules as their valence allows, when the antibody is in great excess. Poliovirus is expected to carry a maximum of 60 molecules of antibody, provided each is attached by a single paratope, as was observed with tobacco mosaic virus in the region of extreme antibody excess (22). That the actual number of antibody molecules per virion was even higher suggests that unspecific binding occurred.…”

Section: Discussionmentioning

confidence: 92%

Relationship between poliovirus neutralization and aggregation

Thomas¹,

Vrijsen²,

Boeyé³

1986

J Virol

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The interaction of mono-and polyclonal neutralizing antibodies with poliovirus was studied. In all cases, neutralization was due to antibody-mediated virus aggregation, and the unpolymerized virions accounted for the residual infectivity. The effect of papain on previously neutralized virus was to deaggregate the virus to fully infective single virions. With some antibodies, the amount of aggregated virus regressed in the region of greatest antibody excess, even though the virus remained fully neutralized. Under these conditions, noninfective, unaggregated immune complexes were formed. A mutant resistant to one of the monoclonal antibodies was selected. The mutant virions were still bound but no longer aggregated or neutralized by the selecting antibodies.

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Section: Discussionmentioning

confidence: 92%

Relationship between poliovirus neutralization and aggregation

Thomas¹,

Vrijsen²,

Boeyé³

1986

J Virol

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“…(2) In the case of peptides and monoclonal antibodies, the antigen valence s = 1. (3) When antibody is injected in small successive amounts, as is the case in a flow, its binding to a multivalent viral antigen tends to be bivalent (Van Regenmortel & Hardie, 1976). In the conditions used for our kinetic runs, it was found that the high concentration of peptide in the dextran was also conducive to bivalent binding (Karlsson et al, 1992).…”

Section: Methodsmentioning

confidence: 93%

Determination of kinetic constants for the interaction between a monoclonal antibody and peptides using surface plasmon resonance

Altschuh

Dubs²,

Weiss³

et al. 1992

Biochemistry

160

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Differences in the affinity of a monoclonal antibody raised against the protein of tobacco mosaic virus for 15 related peptides (residues 134-146) carrying single-residue modifications were investigated using a novel biosensor technology (Pharmacia BIAcore). Analysis of the peptide-antibody interaction in real time allowed fast and reproducible measurements of both association and dissociation rate constants. Out of 15 mutant peptides analyzed, five were not recognized by the antibody at all, and seven were recognized as well as the wild-type peptide. For three of the peptides, the rate constants were different for the mutant and wild-type peptides. The pattern of residue recognition suggests that the epitope is formed by three residues (140, 143, and 144) in a helical conformation that mimics the structure in the protein. Even a minor modification of these residues totally abolishes recognition by the antibody. Modifications of adjacent residues result in small but significant differences in association and/or dissociation rate constants. One of the recognized residues is totally buried in the three-dimensional structure of TMV protein, suggesting that a structural rearrangement next to the helix occurs during protein-antibody interaction.

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“…First, the equilibrium distribution was not reached after 3.5 h. Second, only the virus labelled by antibodies is observed and the buoyant density of the virus- antibody complex is lower than that of the uncomplexed virus. It follows that only 40 out of 780 binding sites present on TMV [15] are occupied by the combination of a specific first antibody, a fluorescent second antibody, and chemical crosslinked between the three components. The profile of the same solution as in Fig.…”

Section: Resolution Of Virus and Free Antibodies In Density Gradient mentioning

confidence: 99%

A fluorescence detection system for the analytical ultracentrifuge and its application to proteins, nucleic acids, and viruses

Schmidt

Rappold

Rosenbaum

et al. 1990

Colloid & Polymer Sci

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A fluorescence detection system was developed for the analytical ultracentrifuge Spinco model E. Fluorescence is excited by a laser beam which is focussed into the cell and illuminates an area with a dimension of 60 ~tm in radial direction. For scanning the laser beam is moved in radial direction. After passing the cell, the laser beam is quenched by a carbon light trap and a set of optical filters. Fluorescence emission intensity is monitored by a photomultiplier located behind the light trap and the set of filters. The sensitivity of the detection system was tested by applying it to the sedimentation analysis of proteins and nucleic acids. Bovine serum albumin (BSA) was covalently labelled with the fluorescence-dye fluorescein-isothiocyanate (FITC), and its sedimentation coefficient could be determined even if BSA was analyzed in a concentration as low as 10-10 M. Nucleic acids were labelled non-covalently by the intercalating dye ethidium bromide. Only 8 ng RNA were needed for the determination of the sedimentation coefficient. The particular advantages of the fluorescence detection system were exploited for the establishment of a new method for quantitative virus detection. To tobacco mosaic virus (TMV) a monoclonal anti-TMV antibody from mouse was bound, and to this a second, anti-mouse antibody that carried the fluorescence-label HTC was attached. Either by UV-irradiation or by incubation with glutaraldehyde, the first antibody was covalently crosslinked to TMV, and the second antibody to the first. In CsC1 density centrifugation with fluorescence detection as little as 3.2 ng virus/80 IA or 6 x 10 s virus particles/ml were recorded in a well expressed band at the corresponding buoyant density. Tenfold lower concentration would result still in a significant band. The sensitivity compares well with those of the most advanced techniques from immunology. Due to the specific labelling of viruses by antibodies it will be possible to carry out quantitative physical characterization of virus containing samples without purifying the virus. Future applications of the fluorescence detection system and of the virus detection technique are discussed.

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Immunochemical studies of tobacco mosaic virus—II

Cited by 41 publications

References 40 publications

Relationship between poliovirus neutralization and aggregation

Relationship between poliovirus neutralization and aggregation

Determination of kinetic constants for the interaction between a monoclonal antibody and peptides using surface plasmon resonance

A fluorescence detection system for the analytical ultracentrifuge and its application to proteins, nucleic acids, and viruses

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