Flow cytometry is a system for sensing cells or particles as they move in a liquid stream through a laser or light beam past a sensing area. The relative light scattering and colour discriminated fluorescence of the particles is measured.
Analysis and differentiation of the cells is based on size and granularity and whether the cell is carrying fluorescent molecules either in the form of antibodies or dyes.
The modern flow cytometer consists of a light source, collection optics, electronics and a computer to translate signals to data. The light source of choice is a laser which emits coherent light at a specified wavelength. Scattered and emitted fluorescent light is collected by two lenses, one set in front of the light source (forward scatter) and one set at right angles (side scatter) and by a series of optics, beam splitters and filters, specific bands of fluorescence can be measured.
Several laser types available that can be used in flow cytometers. They can be gas lasers (e.g. Helium-Neon, Argon, Helium-Cadmium); solid state lasers (e.g. NdYAG); dye lasers or semi-conductors lasers.
The choice of fluorochrome to be used is influenced both by the application and the excitation wavelengths and the availability of an appropriate laser. The following table [PDF - 72KB] lists the major fluorochromes, that are excited by the most commonly used 488nm argon ion laser and emission wavelengths together with their common applications.
It is possible to measure physical characteristics such as cell size, shape and internal complexity and any cell component or function that can be detected by a fluorescent compound can be analysed. The applications of flow cytometry are numerous, and this has led to the widespread use of these instruments in the biological and medical fields.
Numerous books have been published, below are a few:
- Flow Cytometry: A Practical Approach. Edited by MG Ormerod.
(IRL Press, Oxford. 1994. ISBN 0-19 963461-0)
- Practical Flow Cytometry. 3rd Edition. Howard M Shapiro.
(Alan R Liss, Inc. ISBN 0-471-30376-3)
- Flow Cytometry. First Principles. Alice Longobardi Givan.
(Wiley-Liss, New York, 1992. ISBN 0-471-56095-2)
- Handbook of Flow Cytometry Methods. Edited by J Paul Robinson.
(Wiley-Liss, New York, 1993. ISBN 0-471-59634-5).
- Cytometric analysis of cell phenotype and function. Editors DA McCarthy MG Macey (Cambridge University Press, 2001. ISBN0-521-66029-7)
- Flow Cytometry: Principles and Applications. Edited by MG Macey. (Humana Press, 2007. ISBN 978-1-58829-691-7)