Imaging Exocytosis in Retinal Bipolar Cells with TIRF Microscopy

This procedure starts with the dissection of the neural retina of adult goldfish, which is the nervous tissue at the back of the eye. Retinas are placed in an enzymatic solution for 35 minutes, rinsed and ated with a fire polished glass pipette to dissociate cells. Cells are then plated on high refractive glass cover slips and mounted into plastic tissue culture dishes.

Isolated bipolar cells are identified by their distinct morphology to load terminals. We puff on them for 10 seconds with a solution containing FM 1 43 and wait 30 seconds before washing the dye away. After a 30 minute wash cells are patch, clamped and imaged with TIRF microscopy.