TY - JOUR
T1 - Multiphoton-evoked color change of DsRed as an optical highlighter for cellular and subcellular labeling
AU - Marchant, Jonathan S.
AU - Stutzmann, Grace E.
AU - Leissring, Malcolm A.
AU - LaFerla, Frank M.
AU - Parker, Ian
N1 - Funding Information:
Acknowledgments We thank Dr. Charlie Glabe for use of the spectrofluorimeter. This work was supported by NIH grants GM48071 and AG16573. G.E.S. is supported by NRSA training grant AG00096.
PY - 2001
Y1 - 2001
N2 - DsRed, a recently cloned red fluorescent protein, has attracted great interest as an expression tracer and fusion partner for multicolor imaging. We report that three-photon excitation (λ <760 nm) rapidly changes the fluorescence of DsRed from red to green when viewed subsequently by conventional (one-photon) epifluorescence. Mechanistically, three-photon excitation (λ <760 nm) selectively bleaches the mature, red-emitting form of DsRed, thereby enhancing emission from the immature green form through reduction of fluorescence resonance energy transfer (FRET). The "greening" effect occurs in live mammalian cells at the cellular and subcellular levels, and the resultant color change persists for >30 h without affecting cell viability. This technique allows individual cells, organelles, and fusion proteins to be optically marked and has potential utility for studying cell lineage, organelle dynamics, and protein trafficking, as well as for selective retrieval of cells from a population. We describe optimal parameters to induce the color change of DsRed, and demonstrate applications that show the potential of this optical highlighter.
AB - DsRed, a recently cloned red fluorescent protein, has attracted great interest as an expression tracer and fusion partner for multicolor imaging. We report that three-photon excitation (λ <760 nm) rapidly changes the fluorescence of DsRed from red to green when viewed subsequently by conventional (one-photon) epifluorescence. Mechanistically, three-photon excitation (λ <760 nm) selectively bleaches the mature, red-emitting form of DsRed, thereby enhancing emission from the immature green form through reduction of fluorescence resonance energy transfer (FRET). The "greening" effect occurs in live mammalian cells at the cellular and subcellular levels, and the resultant color change persists for >30 h without affecting cell viability. This technique allows individual cells, organelles, and fusion proteins to be optically marked and has potential utility for studying cell lineage, organelle dynamics, and protein trafficking, as well as for selective retrieval of cells from a population. We describe optimal parameters to induce the color change of DsRed, and demonstrate applications that show the potential of this optical highlighter.
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U2 - 10.1038/90249
DO - 10.1038/90249
M3 - Article
C2 - 11433276
AN - SCOPUS:0034930509
SN - 1087-0156
VL - 19
SP - 645
EP - 649
JO - Nature biotechnology
JF - Nature biotechnology
IS - 7
ER -