Figure 2 | Scientific Reports

Figure 2

From: An Integrated Chemical Proteomics Approach for Quantitative Profiling of Intracellular ADP-Ribosylation

Figure 2

Schematic illustrating the experimental workflow for quantitative comparison of metabolic labelling between 2YnAd and 6YnAd. MDA-MB-231 cells were metabolically labelled with 3 different concentrations of 2YnAd and 6YnAd in parallel. After cell lysis and click reaction with a biotin-containing capture reagent, the labelled proteins are affinity enriched on NeutrAvidin-Agarose resins. Stringent washing at this stage removes unmodified proteins from the beads. After on-bead reduction, alkylation and tryptic digestion, the eluted peptides are desalted and each sample is labelled using a unique TMT6plex channel. After quenching the TMT reactions, the 6 samples are mixed together, fractionated and subjected to nanoflow LC-MS/MS analysis. Quantification of reporter ions from the MS/MS spectra provides relative quantification of labelling efficiencies of the two compounds at all three tested concentrations.

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