DNA Switches on the Two-Photon Efficiency of an Ultrabright Triphenylamine Fluorescent Probe Specific of AT Regions

Abstract

We report on the design and synthesis of two-photon fluorescent triphenylamines bearing two or three vinyl branches terminated by a N-methyl benzimidazolium moiety. The new compounds (TP-2Bzim, TP-3Bzim) are light-up fluorescent DNA probes with a long wavelength emission (>580 nm). Compared to their pyridinium models, the TP-Bzim dyes exhibit a remarkable improvement of both their DNA affinity and fluorescence quantum yield, especially for the two-branch derivative (TP-2Bzim: ΦF = 0.54, Ka = 10(7) M(-1)), resulting in a large fluorescence emission turn-on ratio of up to 140. Concomitantly, the two-photon absorption cross-section of TP-2Bzim is dramatically enhanced upon DNA binding (δ = 1080 vs 110 GM for the free form). This effect of the DNA matrix on the nonlinear absorption is uncovered for the first time. This is attributed to a tight fit of the molecule inside the minor groove of AT-rich DNA which induces geometrical rearrangements in the dye ground state as supported by circular dichroism and molecular modeling data. Consequently, TP-2bzim displays an exceptional two-photon molecular brightness (δ×ΦF = 583 GM), a value unrivalled for a small biofluorophore. These properties enable to image nuclear DNA in fixed cells at submicromolar concentration ([TP-2Bzim] = 100 nM) and to visualize ultrabright foci of centromeric AT-rich chromatin. Finally TP-2Bzim exhibits a high photostability, is live-cell permeant, and does not require RNase treatment. This outstanding combination of optical and biological properties makes TP-2Bzim a bioprobe surpassing the best DNA stainers and paves the way for studying further nonlinear optical processes in DNA.

Dynamically Monitoring Cell Viability in a Dual‐Color Mode: Construction of an Aggregation/Monomer‐Based Probe Capable of Reversible Mitochondria‐Nucleus Migration

“…[11][12][13][14][15] Compared with these techniques,fluorescence imaging methods assisted with proper fluorescent probes have received considerable interest in recent years,o wing to the unique advantages including capability of in situ and real-time visualization in live cells,l ow damage to biosamples,a nd permission of dynamically analyzing live samples. [19] Eliseeva et al have presented an ear infrared fluorescent probe for the discrimination of dead cells. [18] Te ulade-Fichou and co-workers have developed two-photon fluorescent probes for the imaging of cell nucleus.…”

Indole-based Cyanine as a Nuclear RNA-Selective Two-Photon Fluorescent Probe for Live Cell Imaging

“…[11][12][13][14] Recently, Teulade-Fichou et al reported a nuclear membrane permeable DNA-selective TPEF probe which bound to duplex DNA in nucleus due to its relatively strong interaction with DNA at the AT-rich region. 15 This probe allowed the investigation of the transcriptional dynamics of the cell nucleus. However, a nuclear RNAselective TPEF probe for live cell imaging is rarely available.…”

Photostability Enhancement of Fluorenone-Based Two-Photon Fluorescent Probes for Cellular Nucleus Monitoring and Imaging

“…20 So far, a number of two-photon probes have been studied for cellular DNA or other organelles targeting and imaging. [21][22][23][24][25] However, studies involving organic small-molecule probes targeting cellular nucleus for the long-term monitoring and tracking are rarely reported. 26,27 Thus, extreme attention should be paid to nucleus monitoring for the enhancement of photostability in biomedical imaging.…”