We present a protocol to evaluate the utility of detergents for purification and delipidation of E. coli membrane proteins. We determine the critical aggregation concentration of detergents. Furthermore, we compare the ability of detergents to extract membrane proteins and to maintain protein-lipid interactions during purification. The protocol describes steps for isolating and delipidating membrane proteins from E. coli membranes by extraction and affinity purification using detergents. The protocol does not enable an absolute quantification of purification outcomes. For complete details on the use and execution of this protocol, please refer to Urner et al.1.