DiSC3(5) fluorescence-based membrane potential study

IG Ines Greco
AE Agnete Plahn Emborg
BJ Bimal Jana
NM Natalia Molchanova
AO Alberto Oddo
PD Peter Damborg
LG Luca Guardabassi
PH Paul R. Hansen
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Freshly sub-cultured E104 cells were labelled with 1 µM 3,3-Dipropylthiadicarbocyanine iodide [DiSC3(5)] (Sigma) in MHB II. The fluorescence spectra of labelled cells were plotted in the LS50B luminescence spectrometer (PerkinElmer) at excitation/emission wavelengths 546 nm/573 nm using time drive application of FLWINLAB software. After reading an initial stable emission spectra of DiSC3(5), labelled cells were treated with B1 or nisin or protonophore CCCP at 1, 1.5 and 1.25 µg/ml, respectively and the change of fluorescence over time was recorded. The increment of DiSC3(5) fluorescence upon addition of antimicrobials or CCCP (FUafter-treatment – FUbefore-treatment) was plotted. This experiment was performed twice and each time with two technical replicates.

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