Total internal reflection fluorescence microscopy (TIRF microscopy) is one example for surface-sensitive microscopy and is therefore only applied to samples that are in contact with a solid, transparent interface (e.g., the mounting glass slide). In TIRF microscopy, the excitation light is (in contrast to conventional fluorescence microscopy) guided to the interface at the angle of total internal reflection. Hence, the excitation light is almost completely reflected, but creates an evanescent (light-)field at the interface, which excites only those parts of the sample that are in close vicinity to the interface (typically the closest 100 to 150 nm). This spatial confinement decreases the noise background (and thus improves the signal-to-noise ratio), but also increases the optical resolution along the optical axis (with respect to usual wide-field or confocal microscopy solutions). Typical applications include imaging of cell structures that are closely located to the cell membrane (e.g., the cytoskeleton) or the transient attachment of biological nanoparticles (such as viruses/virions, vesicles, exosomes) to cell membranes.
The Zeiss Axiovert 200M is an inverted fluorescence microscope equipped with a VisiTron laser-based TIRF extension (excitation at 488 and 561 nm) and a CoolSNAP CCD camera. It provides conventional fluorescence microscopy, FRET and TIRF.