Video-Rate Bioluminescence Imaging of Protein Secretion from a Living Cell

We have established a method of video-rate bioluminescence imaging to visualize exocytotic protein secretion from a single living cell using Gaussia luciferase (GLase) as a reporter protein. The luminescence signals produced by the enzymatic reaction of secreted GLase (luciferase) and coelenterazine (luciferin) are detected with an electron-multiplying charge-coupled device camera. An exocytotic event of protein secretion can be visualized using the protein fused to GLase with a time resolution of 30–500 ms. Signal analyses of the bioluminescence video images reveal a number of exocytotic sites, the frequency of exocytotic events, and the amount of secreted protein on a whole cell. Furthermore, the method can distinguish between secreted and cell surface-bound proteins. Our method is a direct approach to investigate the secretion and localization of proteins on the whole surface of living cells.