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The researcher first employs techniques described in the protocol manual to limit endocytosis and lysosomal targeting that may result in proteolytic cleavage of cytoplasmic domains of multi-membrane spanning proteins. In the second step, the Transmembrane Protein Extraction Reagent is added to dislodge cells from the cell culture dish and then to dissolve the cell membrane. Following membrane dissolution, brief centrifugation is used to remove cellular debris from a supernatant fraction that contains the extracted transmembrane proteins. The supernatant is added to Laemmli Sample Buffer, which is heated, but not boiled, prior to the resolution in SDS-PAGE gels. For Western blots, transfer takes place in a transfer buffer with added SDS and reduced amounts of methanol. The transfer is allowed to proceed approximately 25% longer than typically employed in Western blotting protocols.An easy to follow protocol for cell lysis and preparation of samples for Western Blotting is included with the Transmembrane Protein Extraction Reagent, or can be downloaded in pdf format by clicking the manual tabs.