3D tracking of single molecular motors

We show a new technique that we developed for 3D tracking of single fluorescent molecules inside living cells with nanometer accuracy. To account for the optical sectioning of thick samples we built up a HILO (Highly-Inclined-and-Laminated-Optical-sheet) microscopy system [1], through which we can excite the sample in a widefield configuration by a thin sheet of light, spanning the entire thickness of the cell with a SNR much higher than traditional WF microscopy. Under these illumination conditions, we apply a method capable of reaching nanometer x, y, z localization by exploiting the properties of the point-spread-function of out-of-focus fluorescent probes.