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By Marion G. Macey

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Or, if the erythrocytes have been lysed, by gating on the relevant leukocytes in a display of forward and side light scatter. 6. , the concentration of CD4+ lymphocytes when monitoring the progress of HIV infection) and this can be done in two main ways. , using the volumetric Ortho CytoronAbsolute). Second, the sample can be mixed with a known concentration (as particles per milliliter) of fluorescent microbeads, cells and beads counted, and the cell concentration computed from knowledge of the bead concentration.

4. ISOLATION OF BLOOD MONONUCLEAR CELLS ON FICOLL®–HYPAQUE 1. 3. 2. Centrifuge at 400g for 30 min at room temperature (20–23°C) and allow the rotor to stop with the brake off. 3. The erythrocytes and neutrophils will sediment into/onto the Histopaque while the mononuclear cells form a band between the Histopaque and the plasma, and the platelets will remain largely in the plasma layer. 4. Remove the mononuclear cell band with a pipet, wash the cells by centrifuging twice with 10 mL of medium at 400g for 5 min, and resuspend in 1 mL of medium.

And Sorvajarvi, K. (1992) CD11b is a calcium-dependent epitope in human neutrophils. Biochem. Biophys. Res. Commun. 87, 195–200. 15. McCarthy, D. A. and Macey, M. G. (1993) A simple flow cytometric procedure for the determination of surface antigens on unfixed leucocytes in whole blood. J. Immunol. Methods 163, 155–160. 16. , Lazarus, A. , Semple, J. , and Freedman, J. (1999) Preanalytical requirements for flow cytometric evaluation of platelet activation: choice of anticoagulant. Transfus. Med.

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