Finally, we conducted measurements of MSP nanodiscs to determine the workability of the system. The number of identified peptides of MSP1E3D1 and the sequence coverage map -two crucial metrics in HDX-MS -have demonstrated equal performance compared to control measurements of the free protein, proving the functionality of the developed delipidation setup. Furthermore, we obtained 82.7% sequence coverage of the AcrB membrane protein solubilized in POPC nanodisc, demonstrating the applicability of the system on proteins of interest. This workflow will facilitate membrane protein characterization in HDX-MS to progress our understanding of protein dynamics in lipid environments. Overall, our developments will advance the field of membrane protein structural mass spectrometry, which is now at the point where a lipid milieu must be considered due to its putative relationship with protein structure and function.