This study compares the newly developed Thermo Scientific™ SMART Digest™ kit to classic in-solution protein digestion methods, focusing on protein sequence coverage and identified post-translational modifications (PTMs), including deamidation, oxidation, and glycosylation. A Thermo Scientific™ Acclaim™ VANQUISH™ C18 column with conventional water/acetonitrile-based gradients and the Thermo Scientific™ Vanquish™ Flex UHPLC system were used for separation in combination with the Thermo Scientific™ Q Exactive™ HF Hybrid Quadrupole-Orbitrap™ mass spectrometer.

Peptide mapping is one of the major analyses in biotherapeutics characterization. Here we show peptide mapping with the Thermo Scientific Vanquish Flex UHPLC system. An efficient and reliable separation of peptides obtained from a bovine serum albumin (BSA) digest is done on an Thermos Scientific Acclaim RSLC 120, C18, 2.2 µm Analytical column. Excellent retention time and area precisions allow reliable qualitative and quantitative data interpretation.

The Thermo Scientific UltiMate 3000 RSLC system is applied for the analysis of tryptic digests of proteins. The separation is performed on Thermo Scientific Acclaim RSLC C18 and Acclaim RSLC PolarAdvantage II 2.2µm columns, with UV detection at 210 nm.

In this work, a digestion of bovine serum albumin is resolved using the Thermo Scientific Acclaim RSLC C18 into 127 peaks with an average peak width of 0.27 minutes. For a 90 minute gradient, this works out to a theoretical peak capacity of >300.

The Thermo Scientific HPLC system is applied for the analysis of a b-Lactoglobulin tryptic digest. The separation is performed on a Thermo Scientific Hypersil GOLD HPLC column, with UV detection at 220 nm.