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M. Morimoto and M. Irie
in a molecule [6–14]. So far, based on this methodology, a number of photoswitchable
fluorescent molecules have been synthesized. These molecules are initially fluorescent, while the fluorescence is turned off by excitation energy transfer or electron
transfer when the photochromic unit undergoes photoisomerization.
Super-resolution fluorescence microscopy breaks the diffraction limit of light
and provides fluorescence images with nanometer-scale spatial resolution. The
imaging techniques are usually classified into two categories, coordinate-targeted and
coordinate-stochastic methods. In coordinate-targeted methods, such as reversible
saturable (switchable) optical linear fluorescence transition (RESOLFT) microscopy
[15, 16], the position of the on- and off-states is determined by a doughnut-shaped
pattern of light inducing a turn-on or a turn-off transition in the sample, except at the
center with vanishing intensity. Coordinate-stochastic methods, such as photoactivated localization microscopy (PALM) [17] and stochastic optical reconstruction microscopy (STORM) [18], are based on the detection and localization of
randomly arising fluorescent spots of on-state single molecules. The reversibly photoswitchable fluorescent molecules can be applied to both RESOLFT microscopy and
PALM/STORM. However, the turn-off mode photoswitchable fluorescent molecules
are hardly applicable to PALM/STORM because the quenching of fluorescence, in
most cases, is imperfect and the imaging methods require a dark background to detect
the single-molecule fluorescence. For the application, it is strongly desired to develop
turn-on mode photoswitchable fluorescent molecules, which can be efficiently and
instantaneously activated to be fluorescent upon photoirradiation.
Although several turn-on mode photoactivatable fluorophores have been reported,
the photoactivation quantum yields are relatively low and the reactions are irreversible [19–26]. Recently, a new type of turn-on mode photoswitchable fluorescent
molecules has been developed. They are sulfone derivatives of 1,2-bis(2-alkyl-6-aryl1-benzothiophen-3-yl)perfluorocyclopentene [27, 28]. Although the diarylethene
derivatives are initially non-luminous under excitation with visible light, they are activated to emit strong fluorescence upon irradiation with UV light. The photoswitching
and fluorescent properties are dependent on the substituents of the benzothiophene
rings. In this chapter, we describe photochemical and photophysical properties of the
turn-on mode fluorescent diarylethenes 1–8 (Fig. 33.1 and Table 33.1) and how to
control the photoswitching performance by chemical modifications for the application to super-resolution fluorescence microscopy. Solvatochromism of photoswitchable fluorescence and unprecedented reversible switching with single-wavelength
visible light are also described.
M. Morimoto and M. Irie
in a molecule [6–14]. So far, based on this methodology, a number of photoswitchable
fluorescent molecules have been synthesized. These molecules are initially fluorescent, while the fluorescence is turned off by excitation energy transfer or electron
transfer when the photochromic unit undergoes photoisomerization.
Super-resolution fluorescence microscopy breaks the diffraction limit of light
and provides fluorescence images with nanometer-scale spatial resolution. The
imaging techniques are usually classified into two categories, coordinate-targeted and
coordinate-stochastic methods. In coordinate-targeted methods, such as reversible
saturable (switchable) optical linear fluorescence transition (RESOLFT) microscopy
[15, 16], the position of the on- and off-states is determined by a doughnut-shaped
pattern of light inducing a turn-on or a turn-off transition in the sample, except at the
center with vanishing intensity. Coordinate-stochastic methods, such as photoactivated localization microscopy (PALM) [17] and stochastic optical reconstruction microscopy (STORM) [18], are based on the detection and localization of
randomly arising fluorescent spots of on-state single molecules. The reversibly photoswitchable fluorescent molecules can be applied to both RESOLFT microscopy and
PALM/STORM. However, the turn-off mode photoswitchable fluorescent molecules
are hardly applicable to PALM/STORM because the quenching of fluorescence, in
most cases, is imperfect and the imaging methods require a dark background to detect
the single-molecule fluorescence. For the application, it is strongly desired to develop
turn-on mode photoswitchable fluorescent molecules, which can be efficiently and
instantaneously activated to be fluorescent upon photoirradiation.
Although several turn-on mode photoactivatable fluorophores have been reported,
the photoactivation quantum yields are relatively low and the reactions are irreversible [19–26]. Recently, a new type of turn-on mode photoswitchable fluorescent
molecules has been developed. They are sulfone derivatives of 1,2-bis(2-alkyl-6-aryl1-benzothiophen-3-yl)perfluorocyclopentene [27, 28]. Although the diarylethene
derivatives are initially non-luminous under excitation with visible light, they are activated to emit strong fluorescence upon irradiation with UV light. The photoswitching
and fluorescent properties are dependent on the substituents of the benzothiophene
rings. In this chapter, we describe photochemical and photophysical properties of the
turn-on mode fluorescent diarylethenes 1–8 (Fig. 33.1 and Table 33.1) and how to
control the photoswitching performance by chemical modifications for the application to super-resolution fluorescence microscopy. Solvatochromism of photoswitchable fluorescence and unprecedented reversible switching with single-wavelength
visible light are also described.