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Quantifying Metabolic Activity of Filamentous Fungi Using a Colorimetric XTT Assay
Author(s) -
Moss Bill J.,
Kim Yonghyun,
Nandakumar M. P.,
Marten Mark R.
Publication year - 2008
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1021/bp070334t
Subject(s) - formazan , absorbance , chemistry , biochemistry , dry weight , chromatography , biology , botany
Abstract The filamentous nature and robust cell walls of many fungi render traditional measurements of active biomass (e.g., turbidity, dry cell weight) ineffective for most fungal bioprocesses. To overcome this challenge, an assay for quantification of overall metabolic activity is developed using 2,3‐bis(2‐methoxy‐4‐nitro‐5‐sulfophenly)‐5‐[(phenylamino) carbonyl]‐2 H ‐tetrazolium hydroxide (XTT), which in the presence of active mitochondria is converted to a water‐soluble formazan derivative that absorbs light in the visible spectrum (430‐490 nm). Tests on the model fungus Aspergillus nidulans show that in actively growing cultures XTT absorbance is linearly related to dry cell weight below 0.2 g/kg broth. Validation through growth rate testing shows the developed XTT assay is able to accurately quantify reductions in culture metabolism during damaging physical treatment (heat, high shear, microwaving). Experiments in batch culture demonstrate that the developed XTT assay is capable of reporting on metabolic activity where dry cell weight is not. The developed assay is inexpensive, relatively rapid, and easy to conduct, making it ideally suited for assessment of fungal processes in the biotechnology industry.