Bright field Microscopy
In this modality of microscopy an image of the sample is generated by absorbance of
some of the transmitted light (i.e. light going through the sample) by dense areas
in the sample. A bright field microscopy image typically appears dark on a bright
background, hence the name.
Fluorescence microscopy is used to image fluorescent molecules in a specimen. To that
end, illumination, incident light arriving onto the specimen excites the fluorescent
molecules to emit fluorescent light which is then viewed through the eyepiece. The
incident light is filtered out before it arrives to the eyepieces and therefore the
image appears as bright features on a dark background. The two most common modalities
of fluorescence microscopy are wide-field and confocal scanning.
Wide-field Fluorescence Microscopy
The illumination and fluorescent light cover the whole visual field of the microscope
objective. The image can be viewed through the eyepieces or detected by a camera.
Confocal Scanning Fluorescence Microscopy
This type of fluorescence microscopy is: (1) confocal because it reduces the collection
of light away from the focal plane (i.e. it collects light from optical sections)
and (2) scanning because laser light is scanned through a narrow area of the specimen
to generate fluorescent light; the image is not wide-field because the scanned area
is smaller that the visual field of the microscope objective. In this type of microscopy,
the image cannot be viewed through the eyepieces. Instead, the fluorescent light is
collected by specialized detectors and a dedicated computer program assembles an image
that can be view on a computer monitor and saved as a digital file.