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Figure S1: HCD parameter optimization. All parameters shown were optimized using 1 ug E.coli digest and a 60’ LC gradient time. The evaluation criteria included the number of identified spectra, peptides and proteins.

A) Collision energy (in %)

B) Automatic gain control settings (number of collected precursor ion charges for fragmentation)

C) number of most abundant precursor ions picked for fragmentation in one cycle

D) Signal threshold of precursor ions for triggering an MS/MS spectrum

E) Survey scan start m/z values

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