Flow Cytometry Fluidics

Flow Cytometry Fluidics

The purpose of fluidics system is to transport particles in a fluid stream for laser beam interrogation, with only one cell or particle moving through the laser beam at any given moment. The sample is therefore, injected into a stream of sheath fluid within the flow chamber. The design of the flow chamber causes the sample core to be focused in the center of the sheath fluid. The flow of sheath fluid accelerates the particles and restricts them to the center of the sample core, in a process of hydrodynamic focusing. The calibration of flow cytometers in the clinical laboratory is of primordial importance for the interlaboratory comparison of quantitative and classification results. Brightly fluorescent alignment beads are used to control the overall optical and electronic performance of a flow cytometer. The measurement of a mixture of typically four fluorescent beads with knwon amounts of antibody binding sites or bound antibodies permits the clibration of the fluorescence scale of a flow cytometer in “bound fluorescent antibody molecules”. A non fluorescent bead characterizes the assay noise level of the flow cytometer. When a filter is placed at a 45 degree angle to a light source, light which would have been trasmitted by that filter is still transmitted but light that would have been blocked is reflected (at a 90 degree angle). Used this way, a filter is called a dichroic filter.

References

1. Janeway, C.A., Travers, P., Walport, M., and Capra, J.D. 1999. Immunobiology: The immune system in health and disease. Garland Publishing, 4th ed., New York, USA

2. Delves, P. and Roitt, I. 1999. Encyclopedia of Immunology. Academic Press Inc., 2nd ed., San Diego, USA

3. 1994. Current Protocols in Molecular Biology. Volume 2. John Wiley & Sons Inc., USA

4. Cruse, J. and Lewis R. 1995. Illustrated Dictionary of Immunology. CRC Press Inc., USA

5. http://biology.berkeley.edu/crl/flowcytometry.shtml

6. Lodish, H., Baltimore, D., Berk, A., Zipursky, Matsudaira, and Darnell, J. 2000. Molecular Cell Biology. American Scientific Books, USA

7. www.spectracell.com/jana.html

8. Andrade, W., Johnston, M. and Hay, J.B. 1998. The relationship of blood lymphocytes to the recirclating lymphocyte pool. Blood 91: 1653-1661.