178
edits
Changes
no edit summary
==Developments==
==Developments==
===Confocal microscopy===
===Confocal microscopy===
*The scanning confocal fluorescent microscope is a recent development that uses computer-aided techniques to produce a high-resolution thin optical section of a sample without the need for further, complex sample preparation. Fluorescent images were previously hard to resolve as the picture was subject to glare from planes above and below that in focus, resulting in a blurred image.
*To obtain an image, a laser light is focused on a fine plane within the sample. The resulting fluorescent emission is collected in a photomultiplier tube (PMT) with a confocal aperture. The fluorescence from planes above and below the optical section fails to reach this aperture, resulting in a sharper image.
*The resolution can be further increased by using low-level illumination, meaning the fluorescent dye can only be excited by two photons. When a pulsed laser is used, it only reaches sufficient intensity to excite fluorescence when the beam is focused onto the focal plane of the microscope. This minimises fluorescence to the desired optical section itself.
===Time-lapse video microscopy===
*Sensitive digital video cameras can be used to record the movement of fluorescently tagged molecules, for example to track the movement of such molecules in cell membranes or during interaction between cells.
===Flow cytometry===
*The flow cytometer is an instrument that directs cells in single file through a narrow chamber illuminated by a laser beam. As fluorescently-tagged cells pass through the laser beam, the fluorochrome is excited and emits light that is detected by a PMT-like detector.
*This detector is capable of measuring the strength of emission, so it is possible to sort cells that are negative, slightly positive and highly positive for a specific antigen.