Why is TIRF microscopy useful?
TIRF (Total Internal Reflection Fluorescence) only excites fluorophores on the surface of molecules in the 200 nm layer, so other fluorophores are not excited and do not emit light. As a result background noise is significantly reduced. This improvement means that TIRF has become a leading technique in the study of single molecules.
It is useful for samples with fine structures, thinner than 200 nm and sitting close to the coverslip. This type of imaging is also ideal for studying molecules attached to a surface or on a membrane. In addition it can be employed to study the cytoplasmic zone immediately beneath plasma membranes.