Human islets were dissociated by 10-min incubation in Hanks'-based enzyme-free cell dissociation buffer (GIBCO, Invitrogen) and gentle pipetting to generate small clusters of cells. Dissociated cells were plated onto 13/24-mm sterile coverslips and allowed to recover for 1–2 days. Cells were fixed in 4% paraformaldehyde and permeabilized in 0.1% Triton X-100. Primary cells were blocked in 10% fetal calf serum and subsequently incubated overnight with VPS13C 15-E antibody (Santa Cruz sc-104751, 1:50) with or without anti-insulin antibody (1:200; Dako, Ely, UK) followed by incubations with Alexa 488 and Alexa 568-conjugated secondary antibodies in sequential order. Coverslips were mounted using VectaShield with DAPI and imaged as described elsewhere (42). Samples were illuminated using steady-state 488- and 560-nm laser lines, and emission was collected through ET535/30 and ET620/60 emission filters (Chroma). Images were captured using a Hamamatsu EM CCD digital camera controlled by an Improvision/Nokigawa spinning disc system running Volocit (PerkinElmer, MA) software.