A method of performing proteomic analysis comprising the steps of: placing a single cell sample into a smaller volume nanowell disposed within a chip, the smaller volume nanowell configured to have a diameter < 2 mm and hold liquid volume < 1 μL; placing a peptide mixture into a larger volume booster well, the larger volume booster well configured to have a diameter > 1 mm and hold liquid volume > 1 μL, wherein the diameter of the smaller volume nanowell is less than the diameter of the larger volume booster well; lysing the cell sample; extracting proteins from the cell sample; digesting the proteins into peptides; labeling the peptides and the peptide mixture using tandem mass tag labels; combining the labeled peptides from the nanowells and the booster wells into the booster well to form a mixed sample; separating the mixed sample; and acquiring data from the separated sample using a mass spectrometer, wherein the method further comprises the following steps: the step of setting the automatic gain control levels on the mass spectrometer during MS2 or MS3 data collection > 5E5; setting injection times during MS2 or MS3 data collection > 250 ms; adjusting the automatic gain control levels and injection times based on the booster/sample ratios and the protein abundance in single cells; and wherein separating the mixed sample includes a step of pre-fractionating the mixed sample into multiple fractions. placing a single cell sample into a smaller volume nanowell disposed within a chip, the smaller volume nanowell configured to have a diameter < 2 mm and hold liquid volume < 1 μL; placing a peptide mixture into a larger volume booster well, the larger volume booster well configured to have a diameter > 1 mm and hold liquid volume > 1 μL, wherein the diameter of the smaller volume nanowell is less than the diameter of the larger volume booster well; lysing the cell sample; extracting proteins from the cell sample; digesting the proteins into peptides; labeling the peptides and the peptide mixture using tandem mass tag labels; combining the labeled peptides from the nanowells and the booster wells into the booster well to form a mixed sample; separating the mixed sample; and acquiring data from the separated sample using a mass spectrometer, wherein the method further comprises the following steps: the step of setting the automatic gain control levels on the mass spectrometer during MS2 or MS3 data collection > 5E5; setting injection times during MS2 or MS3 data collection > 250 ms; adjusting the automatic gain control levels and injection times based on the booster/sample ratios and the protein abundance in single cells; and wherein separating the mixed sample includes a step of pre-fractionating the mixed sample into multiple fractions. the step of setting the automatic gain control levels on the mass spectrometer during MS2 or MS3 data collection > 5E5; setting injection times during MS2 or MS3 data collection > 250 ms; adjusting the automatic gain control levels and injection times based on the booster/sample ratios and the protein abundance in single cells; and wherein separating the mixed sample includes a step of pre-fractionating the mixed sample into multiple fractions.