A universal, bottom-up approach analyzes specific IgG tryptic peptides located in the constant region (FC) of the mAB, where the peptides are identical from one compound to another. Published methods utilize either overnight capture, 2 hours to reduce and alkylate the mAB prior to digestion, or digestion with traditional trypsin requiring greater than 4 hours digestion time. To minimize the cost and time for analysis compared to published work, heat stable trypsin and different magnetic beads were evaluated.
To generate the FC specific peptides from a humanized mAB compound for LC-MS/MS quantitation, an immunoprecipitation (IP) procedure with a mAB specific capture antibody was used to selectively capture the mAB of interest from the sample
Evaluate the different capture antibody and different magnetic beads for assay performance
Evaluate the heat stable trypsin from different vendors for assay performance