TIRF Microscopy

Imaging modalities such as epi-fluorescence microscopy can suffer from the presence of out-of-focus light in images.  Confocal microscopy can partially limit this but generally is difficult to perform with live samples due to the amount of light required for illumination and the time required to perform each individual XY scan.  Total internal reflection fluorescence (TIRF) microscopy employs an “evanescent wave” of illumination which only penetrates ~100nm into the sample.  TIRF is a wide-field technique so images can be rapidly acquired on a camera chip, and TIRF is compatible with simultaneous multi-color imaging.  Therefore, TIRF has emerged as the technique of choice for imaging structures and events occurring towards the bottom of live cells (e.g. cell adhesion, endo-/exocytosis and cytoskeletal dynamics).

 

Click to view our J Cell Sci Commentary on TIRF Microscopy in Cell Biology.