Antibodies enzymatic digestion

The MALDI mass spectrum obtained from the peptides resulting from the enzymatic digestion (Asp-N) of the bFGF protein (A) spectrum before and (B) spectrum after immunoprecipitation with an antibody directed against the recombinant protein. The peaks marked correspond to either Cu adducts or doubly charged ions. (C) Schematic representation of the observed peptides. A continuous line corresponds to a peptide that links to the antibody, and a dotted line to a peptide that does not link. Reproduced (modified) from Zhao Y.M., Muir T.W., Kent S.B.FL, Tisher E., Scardina J.M. and Chait B.T., Proc. Natl. Acad. Sci. USA, 93, 4020M024, 199, with permission. 

In the identification step, the protein complex is released from the antibody prior to enzymatic digestion, especially when antibody inunobilization was used, or the antibody-protein complex is digested. The proteins in the complex are then identified by means of PMF and/or PSA using LC-MS or MALDI-MS analysis. 

Amino acid analysis is widely applied in research, clinical facilities, and industry. It is a fundamental technique in biotechnology, used to determine the concentration of peptide solutions, to confirm protein binding in antibody conjugates, and for end-terminal analysis following enzymatic digestion. Clinical applications include diagnosing metabolic disorders in newborns. In industry, it is used for quality control of products ranging from animal feed to infant formula. 

The following protocol is recommended for the majority of antibodies needing enzymatic digestion ... 

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