HeLa cells were lysed and free cysteines were alkylated with an alkyne-tagged light probe (IAA-BnL or IAA-PrL). S-acylation was hydrolysed with hydroxylamine and newly revealed Cys allowed to react with the heavy probe (IAA-BnL or IAA-PrL). Proteins were clicked with azido-biotin, Adadps-biotin, AazoB or ARB, enriched on neutravidin beads, digested with trypsin and desalted. Different optimisations of the protocol were tested: -8 sets of probes / capture reagent -LC-MS/MS optimisation (gradient, FAIMS, etc) Finally, S-acylation was quantified for a biological replicate of HeLa cells.