Allowance for antibody bivalence in the determination of association rate constants by kinetic exclusion assay

Winzor, Donald J. (2013) Allowance for antibody bivalence in the determination of association rate constants by kinetic exclusion assay. Analytical Biochemistry, 441 2: 214-217. doi:10.1016/j.ab.2013.06.020


Author Winzor, Donald J.
Title Allowance for antibody bivalence in the determination of association rate constants by kinetic exclusion assay
Journal name Analytical Biochemistry   Check publisher's open access policy
ISSN 0003-2697
1096-0309
Publication date 2013-10-15
Year available 2013
Sub-type Article (original research)
DOI 10.1016/j.ab.2013.06.020
Open Access Status
Volume 441
Issue 2
Start page 214
End page 217
Total pages 4
Place of publication Philadelphia, PA, United States
Publisher Elsevier
Collection year 2014
Language eng
Formatted abstract
This investigation completes the amendment of theoretical expressions for the characterization of antigen–antibody interactions by kinetic exclusion assay—an endeavor that has been marred by inadequate allowance for the consequences of antibody bivalence in its uptake by the affinity matrix (immobilized antigen) that is used to ascertain the fraction of free antibody sites in a solution with defined total concentrations of antigen and antibody. A simple illustration of reacted site probability considerations in action confirms that the square root of the fluorescence response ratio, RAg/Ro, needs to be taken in order to determine the fraction of unoccupied antibody sites, which is the parameter employed to describe the kinetics of antigen uptake in the mixture of antigen and antibody with defined initial composition. The approximately 2-fold underestimation of the association rate constant (ka) that emanates from the usual practice of omitting the square root factor gives rise to a corresponding overestimate of the equilibrium dissociation constant (Kd)—a situation that is also encountered in the thermodynamic characterization of antigen–antibody interactions by kinetic exclusion assay.
Keyword Antigen–antibody interaction
Antibody bivalence
Association rate constant
Kinetic exclusion assay
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2014 Collection
School of Chemistry and Molecular Biosciences
 
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Created: Wed, 31 Jul 2013, 10:16:41 EST by Mrs Louise Nimwegen on behalf of School of Chemistry & Molecular Biosciences