Antigenic Protein Conjugates and Process for Preparing Same
First Claim
1. A process for preparing an antigenic protein conjugate comprising the steps of:
- (a) contacting an antigenic protein comprising one or more disulfide bonds with a trialkylphosphine under conditions that allow reduction of at least one of said one or more disulfide bonds to provide a reduced antigenic protein comprising at least a pair of sulfhydryl residues, and(b) contacting said reduced antigenic protein with a sulfhydryl reactive reagent comprising a conjugate moiety and a thiol-reactive functionality under conditions that allow reaction of at least one of said sulfhydryl residues with the sulfhydryl reactive reagent to form said antigenic protein conjugate.
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Abstract
An improved process for the preparation of antigenic protein conjugates is provided. The conjugates preferably are formed through reaction with one or more free sulfhydryl groups in the antigenic protein. The process of the present invention preferably employs a trialkylphosphine as the reducing agent and allows for reduction of disulfide bonds in the antigenic protein and conjugation with a conjugate moiety, preferably in a single reaction vessel (i.e. “in situ”) because the process optimally does not require the removal of the reducing agent before subsequent addition of the sulfhydryl reactive agent. Antigenic protein conjugates prepared by the in situ process and their use in diagnostic immunoassays are also provided.
9 Citations
13 Claims
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1. A process for preparing an antigenic protein conjugate comprising the steps of:
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(a) contacting an antigenic protein comprising one or more disulfide bonds with a trialkylphosphine under conditions that allow reduction of at least one of said one or more disulfide bonds to provide a reduced antigenic protein comprising at least a pair of sulfhydryl residues, and (b) contacting said reduced antigenic protein with a sulfhydryl reactive reagent comprising a conjugate moiety and a thiol-reactive functionality under conditions that allow reaction of at least one of said sulfhydryl residues with the sulfhydryl reactive reagent to form said antigenic protein conjugate. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11)
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12. An antigenic protein conjugate prepared by a process comprising the steps of:
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(a) contacting an antigenic protein comprising one or more disulfide bonds with a trialkylphosphine under conditions that allow reduction of at least one of said one or more disulfide bonds to provide a reduced antigenic protein comprising at least a pair of sulfhydryl residues, and (b) contacting said reduced antigenic protein with a sulfhydryl reactive reagent comprising a conjugate moiety and a thiol-reactive functionality under conditions that allow reaction of at least one of said sulfhydryl residues with the sulfhydryl reactive reagent to form said antigenic protein conjugate. - View Dependent Claims (13)
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Specification