Fluorescence microscopy is a kind of indirect imaging technique to enhance the contrast.
The boundary of a biological target is normally not so distinct from the environment, if it is seen as it is. For example, it is difficult to find the transparent cells in the water. There have been a lot of efforts to find ways to enhance the contrast, and fluorescence imaging is the one of powerful and popular imaging methodology.
The fluorophore is bound to a target, and we see the fluorophore through the fluorescene microscope. Staining the target with fluorophore is another important technique as much as imaging technique.
The energy state of fluorophore is excited by the light, and the fluorophore emits the light as this exited energy state is released to the ground state. However the energy level of emitted light is generally lower than the energy level of the excited state, and it makes the wavelength of the emitted light longer than the excitation light. Considering only the visible range of the light, it means the color of the excitation light and the color of the emitted light are different. Because there are a lot of optical components which can filter the specific wavelength range, we can image fluorophore very selectively.
If fluorephor A is bound to a nuclear of a cell and we filter the only emission wavelength range of this fluorophore A, we can see nuclear only. If fluorephor B is bound to a membrane of a cell and we filter the only emission wavelength range of this fluorophore B, we can see membrane only. These selective images are displayed by a different colors on each different channel, and these images from different fluorophore channels are merged together to make one overall integrated image.
Fluorescence image has provided new insight to the biologist. Confocal fluorescence image provides the more higher quality image to the biologist.