In this paper we review the applicability of autofluorescent proteins for single-molecule imaging in biology. The photophysical characteristics of several mutants of the Green Fluorescent Protein (GFP) and those of DsRed are compared and critically discussed for their use in cellular biology. The alternative use of two-photon excitation at the singlemolecule level or Fluorescence Correlation Spectroscopy is envisaged for the study of individual autofluorescent proteins. Single-molecule experiments performed in live cells using eGFP and preferably eYFP fusion proteins are reviewed. Finally, the first use at the single-molecule level of citrine, a more photostable variant of the eYFP is reported when fused to a receptor for neurotransmitter in live cells. To cite this article: L. Cognet et al., C. R. Physique 3 (2002) 645-656. 2002 Académie des sciences/Éditions scientifiques et médicales Elsevier SAS single-molecule / GFP / DsRed / fluorescence microscopy / two-photon excitation / fluorescence correlation spectroscopy Microscopie de fluorescence de protéines autofluorescentes uniques pour la biologie cellulaire Résumé * Correspondence and reprints.