Updated project metadata. To explore the proteomic interactome of Dysferlin in the heart, we used recombinant expression of N-terminally Twin-Strep II-tagged full-length human Dysferlin to establish an in vitro binding assay. Isolated mouse heart membrane fractions were incubated with Dysferlin-coupled beads under four different detergent conditions. Dysferlin pull-down eluates were analyzed by label-free quantification using an ion mobility-enhanced data-independent acquisition (DIA) workflow with alternating low and elevated energy (referred to as UDMSE).