The invention provides bioconjugates of heterocylic compounds such as S-adenosylmethionine and S-adenosylhomocysteine with biotin or digoxigenin. The bioconjugates also include carbon and nitrogen linker moieties of varying length that are used to attach such compounds to biotin or digoxigenin. The conjugates are useful in immunoassays. The invention provides a method for detecting SAM and SAH, comprising the steps of: (a) preparing the following components: (i) bio-conjugates of SAM, SAM analogs or SAH; (ii) an europium, a terbium cryptate or other fluorophore as a donor that has a specific ligand for the tracer in the bio-conjugates of (i); (iii) an acceptor fluorescent dye that has the excitation spectra overlap those of donor's emissions and has an antibody specific for SAM or SAH labeled; (b) addition of the biological fluid containing said SAM or SAH; and (c) spectroscopic measurement of the fluorescence of the donor and the fluorescence of from the acceptor.

Embodiments described herein relate to devices, and methods for quantifying thiol content in a sample containing a mixture of proteins or protein isoforms. The method includes conjugating a portion of the sample with free thiol detection binders, separating the contents in the portion of the sample into separated protein isoforms, detecting fluorescence signals associated with each separated protein isoform, and quantifying, based on the fluorescence signals, a relative amount of free thiol associated with each separated protein isoform. In some instances, the method includes quantifying the amount of each separated protein isoform based on absorbance signals associated with each separated protein isoform. In some instances, the fluorescence and/or absorbance signals associated with protein isoforms conjugated with detection binders can be compared with the corresponding signals associated with unconjugated protein isoforms. In some instances, the method further includes applying a reducing agent and quantifying total-thiol content in the sample.

The present invention relates to a biothiol detecting composition comprising a redox regulation protein, a method for detecting biothiols by using the same, and a biosensor/kit for detecting biothiols. The present invention provides the effect of rapidly measuring free biothiols in body fluids. In addition, relative content ratios and changes of total to free biothiols in body fluids can be detected in real time, which allows biothiols to be available as main indices of diseases through which prediction and warning can be made against various diseases. Further, because various redox stress changes associated with main diseases can be accounted for by variations of biothiols, the present invention can provide important technical, economical, and social values for the investigation of pathogenesis mechanisms and the diagnosis of diseases in the future.

MCE assays and reagents to assess purity and to identify impurities in protein drug product samples are provided. Methods for analyzing analytes in a protein drug sample are provided.

Chemoselective conjugation is achieved through redox reactivity by reacting an N-transfer oxidant with a thioether substrate in a redox reaction in an aqueous environment to form a conjugation product. In embodiments, Redox-Activated Chemical Tagging (ReACT) strategies for methionine-based protein functionalization. Oxaziridine (Ox) compounds serve as oxidant-mediated reagents for direct functionalization by converting methionine to the corresponding sulfimide conjugation product.

Methods of identifying a crosslinking site or a binding site on a protein are described herein. The protein may comprise a binding site in a vicinity of the crosslinking site and identification of the crosslinking site may aid in identifying a location of the binding site in the protein. Methods of identifying a peptide or protein from a complex mixture are also described herein. The invention features a crosslinking agent that is configured to interact with a binding site of a protein and comprises a substituent configured to fragment in tandem mass spectrometry to yield a signature mass fragment.

Compositions and methods for analyzing disulfide bonds are provided. An exemplary method includes preparing peptide standards having no disulfide bonds, scrambled disulfide bond peptide standards, and native disulfide bond peptide standards according to the sequence of the region of the protein drug product that includes the disulfide bond, digesting a sample of protein drug product into peptides, separating the protein drug product peptides, analyzing the protein drug product peptides and the peptide standards, identifying scrambled and native disulfide bond peptides by retention time, and quantifying the level of scrambled disulfide bond peptides.

This present invention provides rapid, reproducible, biomarker-based screening methods for the developmental toxicity testing of compounds. The methods are designed to identify the exposure level at which a test compound perturbs metabolism in a manner predictive of developmental toxicity. In particular, the perturbation of two metabolites, ornithine and cystine, is measured, wherein a ratio of the fold change in ornithine to the fold change in cystine of less than or equal to about 0.88 is indicative of the teratogenicity of a test compound.

Disclosed are methods for preparing and screening for an inhibitor of the activity of a biological molecule having a catalytic or non-catalytic cysteine residue. The methods including preparing a library of candidate inhibitor molecules by conjugating an electrophile to a plurality of drug molecules where the library of candidate inhibitor molecules thus formed react with cysteine residues. The library of candidate inhibitor molecules then may be reacted with the biological molecule to identify those inhibitor molecule that react with the catalytic or non-catalytic cysteine residue of the biological molecule in order to identify an inhibitor of the biological molecule.

A method of identifying subjects with colorectal cancer (CRC) is provided. The method includes obtaining a sample from a subject, determining a level of one or more folate one carbon metabolism (FOCM) metabolites in the sample of the subject, and comparing the level of the FOCM metabolites in the sample of the subject to a level in a non CRC control.