Retention Using Selective Hooks (RUSH) Cargo Sorting Assay for Protein Vesicle Tracking in HeLa Cells

Monitoring vesicle trafficking is an excellent tool for the evaluation of protein dynamics in living cells. Such study is key for the understanding of protein sorting and secretion. Recent developments in microscopy, as well as new methodologies developed to study synchronized trafficking of proteins, allowed a better understanding of signaling, regulation and trafficking dynamics at the secretory pathway. One of the most helpful tools so far developed is the Retention Using Selective Hooks (RUSH) system, a methodology that facilitates the evaluation of synchronized cargo trafficking by monitoring fluorescent vesicles in cells upon biotin addition. Here we present a protocol that allows the quantitative evaluation of protein cargo trafficking at different fixed time points and an analytic approach that enables a better examination of specific cargo trafficking dynamics at the secretory pathway. [GRAPHICS]

Automated summary

Monitoring vesicle trafficking is crucial for evaluating protein dynamics, providing insights into protein sorting and secretion. Recent advancements in microscopy and methodologies for studying synchronized protein trafficking have enhanced understanding of signaling, regulation, and trafficking dynamics in the secretory pathway. The RUSH system is a valuable tool for evaluating synchronized cargo trafficking, allowing for quantitative assessment of protein cargo trafficking at different time points and a detailed examination of specific cargo dynamics at the secretory pathway.