Ultrapurified DsbA and DsbC and methods of making and using the same
AssigneeGenentech, Inc.
InventorsMarc Wong, Liliana T. Yee, Amy Lim, Chris B. Fong
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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.
Claims46
A method for quantifying disulfide oxidoreductase C (Dsb C) in a sample, comprising detecting DsbC in the sample using a detection system and comparing the amount of DsbC detected in the sample with the detection of one or more concentrations of an ultrapure DsbC reference standard, wherein ultrapure DsbC comprises at least about 95% monomeric DsbC.
A method for analyzing a recombinant polypeptide sample for the presence of and/or quantity of DsbC, comprising detecting DsbC in the sample using an immunoassay and comparing the amount of DsbC detected in the sample with the detection of one or more concentrations of an ultrapure DsbC reference standard, wherein the ultrapure DsbC reference standard comprises at least about 95% monomeric DsbC polypeptide.
An immunoassay method for detecting DsbC 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 DsbC with the sample thereby generating a sample-capture antibody combination material;
- (b) contacting a detection antibody that binds DsbC with the sample-capture antibody combination material;
- and
- (c) detecting the 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 DsbC present in the sample based on the level of the detection antibody bound wherein the amount of DsbC present in the sample is determined by comparing the standard titration curve with a standard titration curve generated with an ultrapure DsbC composition, wherein the ultrapure DsbC composition comprises at least about 95% monomeric DsbC.
Dependencies are inferred from the claim text.