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J Antimicrob Chemother. 2005 Jan;55(1):102-5. Epub 2004 Nov 16.

Determination of antifungal drug susceptibilities of Aspergillus species by a fluorescence-based microplate assay.

Author information

1
Program in Infectious Diseases, Fred Hutchinson Cancer Research Center, 1100 Fairview Ave N. D3-100, Seattle, WA 98109, USA. abalajee@fhcrc.org

Abstract

OBJECTIVES:

We have investigated the use of a viability dye, chloromethylfluorescein di-acetate (CMFDA), for antifungal susceptibility testing in a fluorescence microplate (FM) assay format.

METHODS:

For this FM assay, conidia were incubated in increasing concentrations of antifungal drug for 16 h and stained with CMFDA. Fluorescence, measured as mean fluorescence units (MFU) in a fluorescence microplate reader, was graphed relative to that of a drug-free control, and the MIC was defined as the lowest concentration of the drug that resulted in complete reduction (100%) in MFU for amphotericin B, or 90% reduction in MFU for itraconazole and voriconazole. Susceptibilities of 10 clinical isolates of Aspergillus fumigatus, Aspergillus terreus and Aspergillus niger to amphotericin B, itraconazole and voriconazole were tested in a blinded fashion using the FM and the NCCLS methods.

RESULTS AND CONCLUSIONS:

Reproducibility of the FM assay was excellent, and results correlated with those of the NCCLS microdilution method. The FM assay appears to be a rapid, objective method for testing fungal susceptibilities to itraconazole, voriconazole and amphotericin B.

PMID:
15546970
DOI:
10.1093/jac/dkh489
[Indexed for MEDLINE]

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