b'EasyPep mini, maxi, and 96-well plate MS sample prep kitsSimplified sample preparation solutions forHighlights:mass spectrometry Completeincludes preformulated reagents, and an optimized protocol Sample preparation of peptides for MS analysis isfor processing up to 20 samples (mini column), 8 samples (maxi column), complex, with numerous steps and nonstandardor one 96-well plateprotocols, resulting in variable sample quality andCompatiblereagents and protocol designed for compatibility with poor reproducibility. To address these issues, theupstream and downstream processes, including TMT and TMTpro labelingThermo Scientific EasyPep MS Sample Prep KitsFlexiblereagents and protocol have been verified using cells, plasma, have been designed using a standardized workflowand tissue samples for 10 g to 2 mg samplesto improve reproducibility while also saving hands-on and processing time. Time-savingsample processing has been reduced from more than 1 day to less than 4 hours, with improved results (Figures 9 and 10)Protein group identications Unique peptide identicationsTraditional workow4,000 3,459 25,000Peptide19,120Lysis Reduction Alkylation Protein cleanup Protein digestion 3,500 cleanup6,000 50 steps 2,797 20,000 14,848~48 hr2,500 15,000(hr) 0.2 1 1 15 15 0.22,0001,500 10,000EasyPep kit workow 1,000 5,000Figure 9. Comparison of sample preparation procedures 500Reduction/ Peptidefor MS analysis, using a traditional workflow and EasyPep Lysis alkylation Protein digestion cleanup 0 0MS Sample Prep Kits. Compared to a traditional workflow, the EasyPep kit Conventional EasyPep kit Conventional24 steps EasyPep MS Sample Prep Kits produce high-quality peptides inworkow workow24 hr less time, and with half the number of manipulation steps.(hr) 0.1 0.2 13 0.2A Protein grProup identications B Unique peptide identications C TMT labeling eciency TIC areaotein group identications Unique peptide identications4,000 4,000 3,459 3,459 25,000 25,000 120% 7 x 10123,500 3,500 19,120 19,120 100% 99.7 96.6 6 x 1012 5.8 x 1012 5.6 x 10122,797 2,797 20,000 20,0006,000 6,000 14,848 14,848 5 x 101280%2,500 2,500 15,000 15,000 4 x 10122,000 2,000 60%10,000 10,000 3 x 10121,500 1,500 40%1,000 1,000 2 x 10125,000 5,000 20% 12500 500 1 x 100 0 0 0 0% 0EasyPep kitEasyPep kit ConventionalConventional EasyPep kitEasyPep kit ConventionalConventional EasyPep kit Conventional EasyPep kit Conventionalworkow workow workow workow workow workowFigure 10. Comparison of the EasyPep and conventional methods for processing TMT-labeled multiplexed samples. The EasyPep kit resulted in more peptide and protein identifications with high TMT labeling efficiency (N terminus and lysine) compared to a conventional acetone precipitation workflow. TMT labeling eciency TIC areaTIC areaTMT labeling eciencyHow-to video for EasyPep Mini MS Sample Prep Kit:120% 120% 7 x 1012 7 x 101299.7 99.7 12 5.8 x 10125.8 x 1012 5.6 x 10125.6 x 1012100% 100% 96.6 96.6 6 x 10 6 x 101280% 80% 5 x 1012 5 x 10124 x 1012 4 x 1012 Use this rapid mass spec sample prep 60% 60%3 x 1012 3 x 1012 kit for protein extraction, digestion, and 40% 40% 2 x 1012 2 x 1012 peptide cleanup in 24 hours.20% 20% 1 x 1012 1 x 10120% 0% 0 0EasyPep kitEasyPep kit ConventionalConventional EasyPep kitEasyPep kit ConventionalConventionalworkow workow workow workowLearn more at thermofisher.com/easypep96 Mass spectrometry thermofisher.com/proteinbiology'