The development of highly sensitive fluorescent probes in combination with innovative optical techniques is a promising strategy for intravital noninvasive quantitative imaging. Cyanine fluorochromes belong to a superfamily of dyes that have attracted substantial attention in probe design for molecular imaging. We have developed a novel paradigm to introduce a Turn-ON mechanism in cyanine molecules, based on a distinctive change in their π-electrons system. Our new cyanine fluorochrome is synthesized through a simple two-step procedure and has an unprecedented high fluorescence quantum yield of 16% and large extinction coefficient of 52,000 M(-1)cm(-1). The synthetic strategy allows one to prepare probes for various analytes by introducing a specific triggering group on the probe molecule. The probe was equipped with a corresponding trigger and demonstrated efficient imaging of endogenous hydrogen peroxide, produced in an acute lipopolysaccharide-induced inflammation model in mice. This approach provides, for the first time, an available methodology to prepare modular molecular Turn-ON probes that can release an active cyanine fluorophore upon reaction with specific analyte.

The detection of chemical or biological analytes upon molecular reactions relies increasingly on fluorescence methods, and there is a demand for more sensitive, more specific, and more versatile fluorescent molecules. We have designed long wavelength fluorogenic probes with a turn-ON mechanism based on a donor-two-acceptor π-electron system that can undergo an internal charge transfer to form new fluorochromes with longer π-electron systems. Several latent donors and multiple acceptor molecules were incorporated into the probe modular structure to generate versatile dye compounds. This new library of dyes had fluorescence emission in the near-infrared (NIR) region. Computational studies reproduced the observed experimental trends well and suggest factors responsible for high fluorescence of the donor-two-acceptor active form and the low fluorescence observed from the latent form. Confocal images of HeLa cells indicate a lysosomal penetration pathway of a selected dye. The ability of these dyes to emit NIR fluorescence through a turn-ON activation mechanism makes them promising candidate probes for in vivo imaging applications.

Steady-state and ultrafast time-resolved techniques were used to study a newly synthesized photoacid, phenol-carboxyether dipicolinium cyanine dye, QCy9. We found that the excited-state proton transfer (ESPT) to water occurs at the remarkably short time of about 100 fs, k(PT) ≈ 1 × 10(13) s(-1), the fastest rate reported up to now. On the basis of the Förster-cycle, the pK(a)* value is estimated to be -8.5 ± 0.4. In previous studies, we reported the photoacidity of another superphotoacid, the QCy7 for which we found an ESPT rate constant of ~1.25 × 10(12) s(-1), one-eighth that of the QCy9 compound. We found a kinetic isotope effect of the ESPT of about two.

Signal amplification techniques are commonly used to boost diagnostic signals. We have recently developed a new approach for exponential signal amplification obtained through a distinctive dendritic chain reaction. Here we report evaluation of the effect of the self-immolative dendritic amplifier component on the signal amplification. Four dendrons with various numbers of end-units were evaluated for the ability to produce exponential signal amplification through self-immolative disassembly pathways. The dendron composed of a first-generation platform with three end-units exhibited the best characteristics with rapid disassembly rate and good stability under aqueous conditions. This study demonstrates the efficiency of molecules based on dendritic structures.

Steady-state and time-resolved emission spectroscopy techniques were employed to study the excited-state proton transfer (ESPT) to water and D(2)O from QCy7, a recently synthesized near-infrared (NIR)-emissive dye with a fluorescence band maximum at 700 nm. We found that the ESPT rate constant, k(PT), of QCy7 excited from its protonated form, ROH, is ~1.5 × 10(12) s(-1). This is the highest ever reported value in the literature thus far, and it is comparable to the reciprocal of the longest solvation dynamics time component in water, τ(S) = 0.8 ps. We found a kinetic isotope effect (KIE) on the ESPT rate of ~1.7. This value is lower than that of weaker photoacids, which usually have KIE value of ~3, but comparable to the KIE on proton diffusion in water of ~1.45, for which the average time of proton transfer between adjacent water molecules is similar to that of QCy7.