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Fluorescent labeled monoclonal antibodies (mAbs) against CD36 are routinely used as monocyte, erythroid, or platelet markers in clinical cytometry. CD36 has recently been proposed by various authors as a valuable marker helping to enumerate leukocyte's subpopulations by flow cytometry. However, it is known that binding of CD36 may induce platelet activation and formation of platelet's rosettes on leukocytes, resulting in false expression of platelet markers on white blood cells. To study this phenomenon, we have combined classical flow cytometry and a new quantitative flow imaging technique with the ImageStream(®) analyzer. We show that CD36 ligation induces activation of platelets with CD62 expression and their adhesion on leukocytes due to CD62 and CD162 interactions. Preincubation of whole blood samples with either anti-CD62 or anti-CD162 antibodies could prevent formation of these rosettes. Our approach also emphasizes the fact that immunomorphological analysis of cell events with ImageStream technology is a useful tool to validate the specificity of marker's labeling or to elucidate incoherent results obtained with classical flow cytometry. We thus propose to prevent false platelet labeling on leukocytes by preincubation with either anti-CD62 or anti-CD162 antibodies when using CD36 mAbs., (Copyright © 2011 International Society for Advancement of Cytometry.)

3-amino-6-methoxy-9-(2-hydroxyethylamine) acridine (AMHA) is an acridine derivative, which is easily excited in near ultraviolet and which emits a bright green fluorescence. The dye was preferentially incorporated into nucleic structures as attested by microscopic and cytometric analyses after RNase and DNase treatments. The affinity for RNA seemed low and similar to that observed for propidium iodide. AMHA was quickly accumulated in fixed cells, and in appropriate concentrations (10-50 microM) was a DNA- and RNA-specific dye. AMHA probably exhibits an adenine-thymine specificity, as suggested by its quenching after bromodeoxyuridine uptake: the fluorescence quenching was similar to that obtained for Hoechst 33258. After cell treatment by RNase and in the presence of MgCl2, AMHA staining allowed flow cytometric analysis of the cell-cycle distribution. The resulting histograms were similar to those obtained with propidium iodide (CV near 3.5%, and similar cell cycle distribution). Thus, AMHA is a suitable fluorescent dye for efficient analysis of the cell cycle by flow cytometry.