Development of a Kinetic Model To Describe the Effective Rate of Antibody Oxidation by Periodate

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Abstract
The oxidation of antibody carbohydrate residues by periodate is a common approach used for site-specific antibody modification and immobilization. This study sought to develop a general kinetic model that could be used to describe the effective rate of this oxidation for process control. A detailed analysis of previous data collected for rabbit immunoglobulin G in the presence of excess periodate indicated that the reaction followed a pseudo-first-order mechanism in which two general classes of sites were being oxidized. The first class of sites was oxidized fairly rapidly (i.e., within 15−30 min), while the second class of sites reacted over the course of several hours. From these results, an equation was developed that gave a good fit under a variety of reaction conditions to the production of oxidized sites available for coupling with a hydrazide label. On the basis of this equation, data obtained at several periodate concentrations under the same pH and temperature conditions were used to estimate the apparent rate and equilibrium constants for the oxidation of each class of sites. The values obtained by using this approach could be used not only to predict the effective rate of oxidation at other periodate concentrations but also to provide information on the individual steps involved in the oxidation process.