title: Green autofluorescence, a double edged monitoring tool for bacterial growth and activity in micro-plates.
abstract: The intrinsic green autofluorescence of an Escherichia coli culture has long been overlooked and empirically corrected in green fluorescent protein (GFP) reporter experiments. We show here, by using complementary methods of fluorescence analysis and HPLC, that this autofluorescence, principally arise from the secreted flavins in the external media. The cells secrete roughly 10 times more than what they keep inside. We show next that the secreted flavin fluorescence can be used as a complementary method in measuring the cell concentration particularly when the classical method, based on optical density measure, starts to fail. We also demonstrate that the same external flavins limit the dynamical range of GFP quantification and can lead to a false interpretation of lower global dynamic range of expression than what really happens. In the end we evaluate different autofluorescence correction methods to extract the real GFP signal.
authors.affiliation: University Grenoble Alpes/CNRS, LIPhy UMR 5588, Grenoble, F-38041, France.
authors.forename: Irina
authors.initials: I
authors.lastname: Mihalcescu
authors.forename: Mathilde
authors.initials: M
authors.lastname: Van-Melle Gateau
authors.forename: Bernard
authors.initials: B
authors.lastname: Chelli
authors.forename: Corinne
authors.initials: C
authors.lastname: Pinel
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Ravanat
mesh_terms: D001431:Bacteriological Techniques / Q000379:methods*; D002851:Chromatography, High Pressure Liquid; D048168:Escherichia coli K12 / Q000254:growth & development / Q000502:physiology*; D005453:Fluorescence; D049452:Green Fluorescent Proteins / Q000378:metabolism*
chemical_list: D049452:Green Fluorescent Proteins