FLUORESCENCE ASSAY FOR THE DETECTION OF ADHERENT CANDIDA YEASTS TO TARGET-CELLS IN MICROTEST PLATES

We describe an assay based on photometric analysis for the measurement of adherence of Candida species to epithelial target cells (Vero cell line). Adherent Candida cells were detected by staining the cells wit h the fluorescent dye Calcofluor white (CFW), which binds to chitin an d glucan in the yeasts. The tests were performed on microtest plates, which were analysed automatically by fluorescence plate readers. The a ssay is based on the following steps: (i) coating of the microtest pla tes with target cells (e.g. Vero cells); (ii) infection with Candida: (iii) staining of Candida with CFW; (iv) rinsing to remove non-adheren t Candida cells and unbound dye; (v) detection of adherent fluorescent Candida cells. The test was able to detect 4x10(4) cells ml(-1). The standard deviation was +/-8%. Day-to-day variation was +/-10% at most. The adherence of strains of different Candida species was assayed by a standard procedure. The results confirmed the order of adherence, wi th C. albicans ranking first, followed by C. tropicalis, C. parapsilos is and C. glabrata.