Detection and quantification of free sulfhydryls in monoclonal antibodies using maleimide labeling and mass spectrometry.

ABSTRACT: The detection of free sulfhydryls in proteins can reveal incomplete disulfide bond formation, indicate cysteine residues available for conjugation, and offer insights into protein stability and structure. Traditional spectroscopic methods of free sulfhydryl detection, such as Ellman's reagent, generally require a relatively large amount of sample, preventing their use for the analysis of biotherapeutics early in the development cycle. These spectroscopic methods also cannot accurately determine the location of the free sulfhydryl, further limiting their utility. Mass spectrometry was used to detect free sulfhydryl residues in intact proteins after labeling with Maleimide-PEG₂-Biotin. As little as 2% cysteine residues with free sulfhydryls (0.02 mol SH per mol protein) could be d

SUBMITTER: Robotham AC 

PROVIDER: S-EPMC6601545 | biostudies-literature | 2019 May/Jun

REPOSITORIES: biostudies-literature