Confocal laser scanning microscopy

Confocal laser scanning microscopy (CLSM) illuminates a single spot in the XY plane of a sample at a time.  Emitted light is detected by a photomultiplier tube (PMT) after it passes through a “pinhole” that blocks out-of-focus light.  In this way an image is created one point at a time following raster-scanning in XY.  In general it takes ~1s for a conventional CLSM to scan a whole XY field of view (e.g. 512×512 spots), and the standard pinhole size of 1 “Airy Unit” collects all the light emitted at a single spot within an ~1mm depth.