EGFP and DsRed expressing cultures of Escherichia coli imaged by confocal,two-photon and fluorescence lifetime microscopy

The green fluorescent protein (GFP) has become an invaluable marker for mon itoring protein localization and gene expression in vivo. Recently a new re d fluorescent protein (drFP583 or DsRed), isolated from tropical corals, ha s been described [Matz, M.V. et al, (1999) Nature Biotech, 17, 969-973]. Wi th emission maxima at 509 and 583 nm respectively, EGFP and DsRed are suite d for almost crossover free dual color labeling upon simultaneous excitatio n, We imaged mixed populations of Escherichia coli expressing either EGFP o r DsRed by one-photon confocal and by two-photon microscopy, Both excitatio n modes proved to be suitable for imaging cells expressing either of the fl uorescent proteins. DsRed had an extended maturation time and E, coli expre ssing this fluorescent protein were significantly smaller than those expres sing EGFP, In aging bacterial cultures DsRed appeared to aggregate within t he cells, accompanied by a strong reduction in its fluorescence lifetime as determined by fluorescence lifetime imaging microscopy, (C) 2000 Federatio n of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.