Measurement of affinity of viral monoclonal antibodies using fab'-peroxidase conjugate. Influence of antibody concentration on apparent affinity

Azimzadeh, A.; Weiss, Étienne; Regenmortel, M.H.V. Van

doi:10.1016/0161-5890(92)90196-5

Cited by 13 publications

(4 citation statements)

References 20 publications

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“…These results were obtained from binding assays in which free antibody was separated from virusâ ntibody complexes by centrifugation and quantitated by spectrophotometry. More recently, it was shown that the same type of Scatchard analysis could be done at much lower antibody concentration by measuring the amount of free antibody at equilibrium by ELISA titration (Azimzadeh & Van Regenmortel 1991;Azimzadeh et al 1992). It is important to assess if antibodies bind in a monovalent or bivalent manner since the calculated antibody a¤nity may be about two orders of magnitude lower in the case of monovalent binding (Van Regenmortel 1997).…”

Section: Determination Of the Affinity Of Antibodies To Tmvmentioning

confidence: 99%

“…In other virus^antibody systems, the magnitude of the steric e¡ect and the error in a¤nity-constant determination is likely to be even smaller than with TMV, since the protein subunits of most viruses tend to be larger than 17 500 Da and the u/s ratio will thus be smaller. Detailed experimental procedures for measuring the a¤nity constant of TMV antibodies have been described in several reviews (Azimzadeh et al 1992;Pellequer & Van Regenmortel 1993a,b;Saunal et al 1997;Van Regenmortel 1997).…”

Section: The Antigenic Valency Of Tmvmentioning

confidence: 99%

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The antigenicity of tobacco mosaic virus

Regenmortel

1999

Phil. Trans. R. Soc. Lond. B

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The antigenic properties of the tobacco mosaic virus (TMV) have been studied extensively for more than 50 years. Distinct antigenic determinants called neotopes and cryptotopes have been identi¢ed at the surface of intact virions and dissociated coat protein subunits, respectively, indicating that the quaternary structure of the virus in£uences the antigenic properties. A correlation has been found to exist between the location of seven to ten residue-long continuous epitopes in the TMV coat protein and the degree of segmental mobility along the polypeptide chain. Immunoelectron microscopy, using antibodies speci¢c for the bottom surface of the protein subunit, showed that these antibodies reacted with both ends of the stacked-disk aggregates of viral protein. This ¢nding indicates that the stacked disks are bipolar and cannot be converted directly into helical viral rods as has been previously assumed. TMV epitopes have been mapped at the surface of coat protein subunits using biosensor technology. The ability of certain monoclonal antibodies to block the cotranslational disassembly of virions during the infection process was found to be linked to the precise location of their complementary epitopes and not to their binding a¤nity. Such blocking antibodies, which act by sterically preventing the interaction between virions and ribosomes may, when expressed in plants, be useful for controlling virus infection.

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Section: Determination Of the Affinity Of Antibodies To Tmvmentioning

confidence: 99%

Section: The Antigenic Valency Of Tmvmentioning

confidence: 99%

The antigenicity of tobacco mosaic virus

Regenmortel

1999

Phil. Trans. R. Soc. Lond. B

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“…Competitive or blocking ELISA formats using the inhibition between protein–protein or protein–antibody interactions are well-known methods 6 , 65 , 66 . Theoretically, the DENV-serotype-specific IgG detection by the competitive ELISA format is also possible by using an antibody–antibody sandwich system, even if these antibodies have no specificity to each DEN-NS1 serotype (Fig.…”

Section: Resultsmentioning

confidence: 99%

Competitive ELISA for a serologic test to detect dengue serotype-specific anti-NS1 IgGs using high-affinity UB-DNA aptamers

Matsunaga

Kimoto

Lim

et al. 2021

Sci Rep

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Serologic tests to detect specific IgGs to antigens related to viral infections are urgently needed for diagnostics and therapeutics. We present a diagnostic method for serotype-specific IgG identification of dengue infection by a competitive enzyme-linked immunosorbent assay (ELISA), using high-affinity unnatural-base-containing DNA (UB-DNA) aptamers that recognize the four categorized serotypes. Using UB-DNA aptamers specific to each serotype of dengue NS1 proteins (DEN-NS1), we developed our aptamer–antibody sandwich ELISA for dengue diagnostics. Furthermore, IgGs highly specific to DEN-NS1 inhibited the serotype-specific NS1 detection, inspiring us to develop the competitive ELISA format for dengue serotype-specific IgG detection. Blood samples from Singaporean patients with primary or secondary dengue infections confirmed the highly specific IgG detection of this format, and the IgG production initially reflected the serotype of the past infection, rather than the recent infection. Using this dengue competitive ELISA format, cross-reactivity tests of 21 plasma samples from Singaporean Zika virus-infected patients revealed two distinct patterns: 8 lacked cross-reactivity, and 13 were positive with unique dengue serotype specificities, indicating previous dengue infection. This antigen-detection ELISA and antibody-detection competitive ELISA combination using the UB-DNA aptamers identifies both past and current viral infections and will facilitate specific medical care and vaccine development for infectious diseases.

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“…Binding data analysis and linearly converted graphs of the mass action law can be used to compute antibody IRF [16,17]. The most popular of these techniques is a linear variant of the Lineweaver-Burk plot [18].…”

Section: Determination Of Mab Immunoreactive Fractionmentioning

confidence: 99%

An Overview of ELISA-Based Initial Velocity Methods to Measure the Immunoreactive Fraction, Association Rate, And Equilibrium Constants of Monoclonal Antibodies

Toraño¹,

Moreno²,

Infantes³

et al. 2022

Asian J Complement Altern Med

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Three biochemical parameters—the on-rate (k+1), the dissociation rate (k-1), and the equilibrium (Ka) constants—describe the antibody interaction with a ligand. In addition, the antibody immune reactive fraction (IRF) is important because it affects k+1 and Ka constant values. The on-rate and dissociation constants are kinetic, but the equilibrium constant is a thermodynamic parameter, and the methods for their determination are different. Among the available methods to determine these parameters, only surface plasmon resonance (SPR)-based techniques allow calculation of the three antibody constants in a single experiment. However, conventional immunoassays are also suitable to determine antibody constants. Here, we describe some enzyme immunoassays (ELISA) methods developed in our laboratory to, i) quantitate mAb in culture supernatants from the initial velocity of the progress curve, ii) determine mAb IRF by a two-step ELISA method, iii) calculate mAb on-rate constant by kinetic ELISA based on time-course data of ligand binding to capture mAb, and, iv) describe a competitive-inhibition ELISA to measure the inhibition constant (Ki) of mAb binding to plate-bound ligand, in the presence of competing concentrations of mAb and soluble ligand.

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Measurement of affinity of viral monoclonal antibodies using fab'-peroxidase conjugate. Influence of antibody concentration on apparent affinity

Cited by 13 publications

References 20 publications

The antigenicity of tobacco mosaic virus

The antigenicity of tobacco mosaic virus

Competitive ELISA for a serologic test to detect dengue serotype-specific anti-NS1 IgGs using high-affinity UB-DNA aptamers

An Overview of ELISA-Based Initial Velocity Methods to Measure the Immunoreactive Fraction, Association Rate, And Equilibrium Constants of Monoclonal Antibodies

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