Protocol for the detection of large dense-core vesicle exocytosis using an automated image-processing algorithm

Investigating exocytosis in human pancreatic islet cells is challenging due to small vesicle size and variable imaging parameters. Here, we present a protocol to detect and analyze exocytosis with an image-processing algorithm using Lagrangian particle tracking. We describe steps for sample preparation, total internal reflection fluorescence (TIRF) microscopy imaging, and computational analysis. The algorithm is validated by mathematical models, TetraSpeck beads, and cell images. Its applicability to other cellular processes and its handling of large sets of data make it useful for high-throughput microscopy research. For complete details on the use and execution of this protocol, please refer to Makam et al.¹.