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. Author manuscript; available in PMC: 2021 Dec 15.
Published in final edited form as: Anal Chem. 2020 Nov 24;92(24):15726–15735. doi: 10.1021/acs.analchem.0c02533

Figure 1.

Figure 1.

Cloud point extraction process flow and SEC fractionation of endogenous membrane proteins from HEK293T cells. (a) Illustration of the cloud point extraction for the enrichment of membrane proteins. (b) A representative chromatogram of membrane proteins separated by SEC with albumin (66 kDa) and ubiquitin (8 kDa) serving as elution reference standards. SEC was performed using a 1000Å PolyHYDROXYETHYL A (PolyHEA) column and a hydrophobic mobile phase consisting of 40% isopropanol (IPA) and 1% formic acid (FA). (c) SDS-PAGE analysis with coomassie blue visualization of SEC fractions 1–4 (F1, F2, F3, F4). (d) RPLC-MS analysis of SEC fractions. The average proteoform mass (error bars represent standard error of the mean) and the number of proteoforms detected were plotted for each fraction. (e) Representative deconvoluted (low resolution, maximum entropy)48 mass spectra from each fraction.