Fluorescence microscopy differs from other microscopy methods in a variety of ways.; Normal brightfield microscopy in a high power microscope takes light and passes it through a thin specimen on a glass microscope slide.; Phase contrast microscopy begins with normal light, then shifts it out of phase and this phase shift is translated into a difference in viewing intensity.; Darkfield microscopy is an illumination technique where the biological specimen is illuminated from the sides instead of from the bottom.
Fluorescence microscopy differs from all these techniques in that the visible light in the microscope eyepieces is not the original light emitted by the light source.; The light you see is actually light that has fluoresced from the fluorescing microscope specimen.; A high intensity light source is used.; This light is passed through an dichroic filter cube containing a fluorescence bandpass excitation filter.; Only specific wavelengths of light are allowed to pass and reach the fluorescence specimen.; After the incident filtered light reaches the specimen, it is no longer used, and any amount reflecting back into the microscope objective to the dichroic filter is filtered out by the emission filter.; The specimen fluoresces and it is this fluorescing light that passes back through the fluorescence emission filter and goes to the microscope eyepieces to provide a fluorescence image of the specimen.