Long-Wavelength Photoconvertible Dimeric BODIPYs for Super-Resolution Single-Molecule Localization Imaging in Near-Infrared Emission

Abstract

Sulfoxide-bridged dimeric BODIPYs were developed as a new class of long-wavelength photoconvertible fluorophores. Upon visible-light irradiation, a sulfoxide moiety was released to generate the corresponding α,α-directly linked dimeric BODIPYs. The extrusion of SO from sulfoxides was mainly through an intramolecular fashion involving reactive triplet states. By this photoconversion, not only were more than 100 nm red shifts of absorption and emission maxima (up to 648/714 nm) achieved but also stable products with bright fluorescence were produced with high efficiency. The combination of photoactivation and red-shifted excitation/emission offered optimal contrast and eliminated the interference from biological autofluorescence. More importantly, the in situ products of these visible-light-induced reactions demonstrated ideal single-molecule fluorescence properties in the near-infrared region. Therefore, this new photoconversion could be a powerful photoactivation method achieving superresolution single-molecule localization imaging in a living cell without using UV illumination and cell-toxic additives.

Photoactivatable Fluorophores for Bioimaging Applications

“…Negligible influence of irradiation on live cells was observed only at wavelengths longer than 500 nm with the typical power densities of fluorescence photoactivation. Nonetheless, only a few examples of fluorescence photoactivation with green or red light in live cells have been reported so far. − Molecular constructs capable of activating their fluorescence in response to green or even red light, possibly without the participation of triplet states, must be developed to avoid the detrimental effects that the short λ Ac of current PAFs have on biological samples.…”

Photoactivatable BODIPYs for Live-Cell PALM

“…Thus, PAFs, operating on the basis photoinduced bathochromic absorption shifts rather than intramolecular quenching, should ideally be the photoactivatable probes of choice for PALM. Nonetheless, both photoactivation mechanisms have been used so far to reconstruct PALM images of live cells with photoactivatable cyanine [ 40 , 41 ], dihydrofuran [ 42 ], oxime [ 43 ], rhodamine [ 44 , 45 , 46 , 47 , 48 ], thioimide [ 49 ] and BODIPY [ 50 , 51 , 52 , 53 ] derivatives. The latter family of PAFs is the focus of this review.…”

“…The fluorescence of BODIPY fluorophores can be photoactivated with the aid of compatible photocages ( Figure 2 a) relying on either intramolecular quenching ( Figure 3 a) or bathochromic shifts ( Figure 3 b) [ 35 ]. Indeed, several photoactivatable BODIPYs have already been designed around these operating principles and reported in the literature [ 39 , 50 , 51 , 52 , 53 , 54 , 55 , 56 , 57 , 58 , 59 , 60 , 61 , 62 , 63 , 64 ]. We implemented both photoactivation mechanisms, relying on the established photochemistry of ortho -nitrobenzyl (ONB) photocages [ 65 ].…”

“…In fact, only diffraction-limited images were reported in these seminal studies. The photochemical and photophysical properties of compound 24 ( Figure 13 ) overcome some of these limitations; however, this PAF requires green, rather than red, light for photoactivation [ 53 ]. This molecule incorporates two BODIPY components connected through a sulfinyl group.…”

“… Diffraction-limited ( a ) and PALM ( b ) images of the lysosomes of live RAW264.7 cells labeled with 24 [adapted with permission from Ref. [ 53 ]]. …”