[Microbeads and flow cytometry: how and why put the "-metry" in immuno-cytometry?].
In current practice of immunophenotyping, flow cytometers are mainly used as cell counters. These instruments which measure fluorescence intensity on each individual cell with high sensitivity can also count molecules on cells. "Putting the - metry into cytofluorometry" means measuring the expression level of molecules of biological interest on (in) target cells. Candidate molecules are numerous due to the many membrane receptors which biological function depends on the number of accessible molecules. Any modulation of the expression level brings valuable information to the biologist, beit associated to physiological differentiation, pathological state or therapeutic intervention. In order to take the best from such measurements in clinical biology, full reliability is mandatory in terms of time-to-time, platform-to-platform and lab- to-lab reproducibility. One should ban "arbitrary units" as currently provided by the instruments to switch to real units. This paper tends to review the most important features conditioning a reliable quantitation of cellular antigens, with data expressed in terms of number of molecules per cell. In addition to staining tools (antibody-based reagents) and data treatment softwares, calibration tools are mandatory. They are not all equivalent. Fluorescent beads with internal fluorescence are very useful for instrument quality-control but they can not interchange with immunological calibration systems which measure the number of antibody molecules that can bind onto the cells. Numerous application examples illustrate the interest of this "quantitative" dimension of Immunocytometry. Generalization of this approach would help getting a wider access to clinical biology.
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D003584:Cytological Techniques; D005434:Flow Cytometry / Q000295:instrumentation* / Q000379:methods*; D007158:Immunologic Techniques; D008863:Microspheres