Sensitivity of second harmonic generation from styryl dyes to transmembrane potential.
In this article we present results from the simultaneous nonlinear (second harmonic generation and two-photon excitation fluorescence) imaging and voltage clamping of living cells. Specifically, we determine the sensitivity to transmembrane potential of second harmonic generation by ANEP-chromophore styryl dyes as a function of excitation wavelength and dye structure. We have measured second harmonic sensitivities of up to 43% per 100 mV, more than a factor of four better than the nominal voltage sensitivity of the dyes under "one-photon" fluorescence. We find a dependence of voltage sensitivity on excitation wavelength that is consistent with a two-photon resonance, and there is a significant dependence of voltage sensitivity on the structure of the nonchromophore portion of the dyes.
Millard
Andrew C
AC
Department of Physiology and Center for Biomedical Imaging Technology, University of Connecticut Health Center, Farmington, Connecticut 06030-1507, USA.
Jin
Lei
L
Wei
Mei-De
MD
Wuskell
Joseph P
JP
Lewis
Aaron
A
Loew
Leslie M
LM
D000818:Animals; D045744:Cell Line, Tumor; D002934:Cinnamates / Q000378:metabolism; D005456:Fluorescent Dyes; D007834:Lasers; D008564:Membrane Potentials; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000379:methods*; D009447:Neuroblastoma / Q000473:pathology* / Q000503:physiopathology*; D017711:Nonlinear Dynamics; D018408:Patch-Clamp Techniques / Q000379:methods*; D011726:Pyridinium Compounds; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013194:Staining and Labeling / Q000379:methods*
D002934:Cinnamates; D005456:Fluorescent Dyes; D011726:Pyridinium Compounds; C029010:cinnamic acid; C050019:1-(3-sulfonatopropyl)-4-(beta)(2-(di-n-butylamino)-6-naphthylvinyl)pyridinium betaine