Ultrapurified DsbA and DsbC and methods of making and using the same
AssigneeGenentech, Inc.
InventorsMarc Wong, Liliana T. Yee, Amy Lim, Chris B. Fong
Life of this patent
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Abstract
The present invention provides methods for producing disulfide oxidoreductase A (DsbA) and disulfide oxidoreductase C (DsbC) polypeptides at very high levels of purity. Also provided are ultrapure DsbA and DsbC and methods of using same, e.g., for use in immunoassays to show removal of DsbA and DsbC from biologics produced in bacteria.
Claims41
A method for quantifying disulfide oxidoreductase A (DsbA) in a sample, comprising detecting DsbA in the sample using a detection system and comparing the amount of DsbA detected in the sample with the detection of one or more concentrations of an ultrapure DsbA reference standard, wherein ultrapure DsbA comprises at least about 95% monomeric DsbA.
A method for analyzing a recombinant polypeptide sample for the presence of and/or quantity of DsbA, comprising detecting DsbA in the sample using an immunoassay and comparing the amount of DsbA detected in the sample with the detection of one or more concentrations of an ultrapure DsbA reference standard, wherein the immunoassay comprises antibodies that specifically bind ultrapure DsbA which are used as capture antibodies and/or detection antibodies, wherein ultrapure DsbA comprises at least about 95% monomeric DsbA.
An immunoassay method for detecting DsbA in a sample, wherein the sample is obtained from a recombinant polypeptide preparation or a host cell line, the method comprising:
- (a) contacting a capture antibody that binds DsbA with the sample thereby generating a sample-capture antibody combination material;
- (b) contacting a detection antibody that binds DsbA with the sample-capture antibody combination material;
- (c) detecting the detection antibody bound to the sample-capture antibody combination material;
- (d) quantifying the level of the detection antibody bound using a standard titration curve and calculating an amount of DsbA present in the sample based on the level of the detection antibody bound wherein the amount of DsbA present in the sample is determined by comparing the standard titration curve with a standard titration curve generated with an ultrapure DsbA composition, wherein the ultrapure DsbA composition comprises at least about 95% monomeric DsbA.
Dependencies are inferred from the claim text.