G Cozon1, J Ferrándiz, D Peyramond, J Brunet. 1. Unité d'Immunologie, Hôpital de la Croix-Rousse, Lyon Cedex 04, 69317, France. gcozon@rockefeller.univ-lyon1.fr
Abstract
BACKGROUND: human basophils release mediators of allergy after cross-linking of IgE receptors by allergens. Specific activation of basophils is detectable through flow cytometry (FCM) using an anti-CD63 fluorescein-conjugated monoclonal antibody. OBJECTIVES: this study evaluate the detection of activated basophils by FCM in routine diagnosis of atopic diseases as regard to skin prick tests and specific immunoglobulin E antibodies. METHODS AND RESULTS: whole blood from twenty patients suspected of atopy was preincubated with interleukin-3 (IL-3), then incubated with specific allergens. After staining using anti-CD63 antibodies, activated basophils were detected through FCM. IL-3-preincubation increases the spontaneous expression of CD63 even at low concentrations (0.1 ng/ml) on the basophils of 2 patients out of 20. The sensitivity and specificity of FCM were respectively 0.56 +/- 0.17 (m +/- SD) and 1.0 +/- 0.0 for the detection of dust mite-activated basophils without IL-3 preincubation, and 0.73 +/- 0.13 and 1.0 +/- 0.0 for the detention of grass pollen-activated basophils. IL-3-preincubation increased the sensitivity in a dose-dependent manner but decreased the specificity fo FCM for detecting dust mite hypersensitivity. CONCLUSION: this method allow for rapid and easy detection of activated basophils from whole blood, and could be of interest for detecting allergies to non-conventional allergens such as pharmaceutical drugs.
BACKGROUND:human basophils release mediators of allergy after cross-linking of IgE receptors by allergens. Specific activation of basophils is detectable through flow cytometry (FCM) using an anti-CD63fluorescein-conjugated monoclonal antibody. OBJECTIVES: this study evaluate the detection of activated basophils by FCM in routine diagnosis of atopic diseases as regard to skin prick tests and specific immunoglobulin E antibodies. METHODS AND RESULTS: whole blood from twenty patients suspected of atopy was preincubated with interleukin-3 (IL-3), then incubated with specific allergens. After staining using anti-CD63 antibodies, activated basophils were detected through FCM. IL-3-preincubation increases the spontaneous expression of CD63 even at low concentrations (0.1 ng/ml) on the basophils of 2 patients out of 20. The sensitivity and specificity of FCM were respectively 0.56 +/- 0.17 (m +/- SD) and 1.0 +/- 0.0 for the detection of dust mite-activated basophils without IL-3 preincubation, and 0.73 +/- 0.13 and 1.0 +/- 0.0 for the detention of grass pollen-activated basophils. IL-3-preincubation increased the sensitivity in a dose-dependent manner but decreased the specificity fo FCM for detecting dust mite hypersensitivity. CONCLUSION: this method allow for rapid and easy detection of activated basophils from whole blood, and could be of interest for detecting allergies to non-conventional allergens such as pharmaceutical drugs.