A cysteine-selective fluorescent probe for the cellular detection of cysteine (original) (raw)
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pH-Dependent Fluorescent Probe That Can Be Tuned for Cysteine or Homocysteine
Organic letters, 2017
The very close structural similarities between cysteine and homocysteine present a great challenge to achieve their selective detection using regular fluorescent probes, limiting the biological and pathological studies of these two amino thiols. A coumarin-based fluorescent probe was designed featuring pH-promoted distinct turn-on followed by ratiometric fluorescence responses for Cys and turn-on fluorescence response for Hcy through two different reaction paths. These specific responses demonstrate the activity differences between Cys and Hcy qualitatively for the first time. The probe could also be used for Cys and Hcy imaging in living cells.
Tetrahedron Letters, 2011
Two fluorescence probes for the detection of cysteine (Cys), glutathione (GSH) and other biothiols, such as homocysteine (Hcy) and cysteinyl-glycine (Cys-Gly), were developed. These molecular probes are coumarin-based derivatives containing a chalcone-like moiety that reacts with biothiols through a Michael addition reaction, leading to strong fluorescence enhancements. The reactivity of the tested biothiols toward both probes (ChC1 and ChC2) follows the order Cys > GSH > Hcy > Cys-Gly, ChC1 being less reactive than ChC2. Possible interference with other amino acids was assessed. ChC1 and ChC2 display a highly selective fluorescence enhancement with thiols, allowing these probes to be used for fluorimetric thiol determination in SH-SY5Y cells.
Sensors and Actuators B: Chemical, 2014
NBD-chloride is widely used as an efficient probe for selective labelling of thiols in proteins due to formation sulfur-substituted NBD under physiological conditions. Selective conjugation involving thiolate group is favoured over amines of proteins because, amino-substituted NBD can be formed only under more basic and elevated temperature conditions. Sulfur-substituted NBDs generally display weak fluorescence properties compared to amino-substituted derivatives. However, a sulfur-substituted NBD can be converted to corresponding amino-substituted derivative via S-N Smiles rearrangement. Theoretical calculations predicted off-fluorescence state for either the probe or the sulfur-substituted NBD formed upon addition of cysteine. On-fluorescence state was predicted for corresponding amino-substituted NBD derivative. Based on UV-vis and fluorescence spectroscopic studies, most efficient rearrangement was observed for cysteine. The rearrangement was relatively slower for homocysteine and not feasible for glutathione. Detection of cysteine and homocysteine by the probe resulted in 1599-and 760-fold off-on fluorescence enhancements, respectively. Sensing of cysteine by the probe provided a detection limit of 2.0 × 10 −8 M. The sensing of intracellular cysteine by the probe was also demonstrated by live cell imaging.
Sensors and Actuators B: Chemical, 2014
A novel colorimetric fluorescent chemodosimeter (1) based on diketopyrrolopyrrole (DPP) and indanedione for the selective detection of cysteine (Cys) over glutathione (GSH) was synthesized, which was involved by the conjugate addition of Cys to ␣,-unsaturated ketones. The probe featured a fast response (a response time less than 2 min), excitation and emission in the visible region, dual-channel and high selectivity. Addition of Cys in PBS (pH = 7.4) to 1 in THF resulted in a rapid color change from purple to yellow together with appearance of a new absorption peak at 480 nm, while other amino acids did not induce any significant color change. Meanwhile, the Michael addition of Cys to 1 elicited 4.2-fold PL enhancement at 505 nm, which resulted in emission color change from deep red to yellow. Furthermore, 1 could be used as a fluorescent probe for detection Cys 34 within BSA. In addition, the cellular imaging of human adult skin fibroblast cells indicated red fluorescence of 1 was present in the cytoplasm. The CCK-8 assay showed that the cytotoxicity of 1 was low.
Chemical Science, 2012
General Information: Unless otherwise noted, materials were obtained from Aldrich and were used without further purification. 1 H NMR and 13 C NMR spectra were recorded on Brucker AM-300 spectrometers. 1 H NMR and 13 C NMR in CDCl 3 were measured on a Bruker AM-300 spectrometer with tetramethylsilane (TMS) as internal standard. Mass spectra were obtained using a JMS-HX 110A/110A Tandem Mass Spectrometer (JEOL). UV−vis spectra were obtained using a Scinco 3000 spectrophotometer (1 cm quartz cell) at 25 °C. Fluorescence spectra were recorded on RF-5301/PC (Shimada) fluorescence spectrophotometer (1 cm quartz cell) at 25 °C. Deionized water was used to prepare all aqueous solutions. Methods for cell culture and fluorescent imaging Human breast carcinoma (MCF-7) cells were seeded on 18 mm-glass coverslips (Marienfeld, Lauda-Koenigshofen, Germany) at density 2×10 5 cells and cultured in McCoy's 5a media with 10% bovine calf serum and 26 mM sodium carbonate at 37 ºC in a humidified incubator containing 5% CO 2 and 95% air. In order to induce oxidative stress, cells were rinsed three times with phosphate-buffered saline (PBS) and incubated in glucose-free DMEM (Dulbecco's Modified Eagle Media) without antibiotics and bovine calf serum for 2 h. 17 After the incubation, MCF-7 cells were rinsed with PBS and then incubated with 5 M of CyAc for 30 min at RT. The treated cells were washed with PBS and mounted onto a glass slide with ClearMount aqueous mounting medium (Invitrogen). To visualize the NIR fluorescence a zenon lamp (Hamamatsu, Shizuoka, Japan; 75 watt) and cy7 filter cube (Semrock, Rochester, NY; Ex. 660-750 nm/Em. 760-855 nm) was used in comparison with Hg 2+ lamp (Nikon; 100 watt) and Nikon filter cube (G-2A; Ex. 510-560 nm/Em. 590 nm) for 535 nm absorption peak of CyAc. Fluorescent images of the mounted cells were obtained by using an inverted microscope (Nikon Eclipse TE2000-U) at various magnifications (100 to 200 ).
Analytical chemistry, 2018
The development of novel fluorescent probes for monitoring the concentration of various biomolecules in living systems has great potential for eventual early diagnosis and disease intervention. Selective detection of competitive species in biological systems is a great challenge for the design and development of fluorescent probes. To improve on the design of fluorescent coumarin-based biothiol sensing technologies, we have developed herein an enhanced dual emission doubly activated system (DACP-1 and the closely related DACP-2) for the selective detection of glutathione (GSH) through the use of one optical channel and the detection of cysteine (Cys) by another channel. A phenylselenium group present at the 4-position completely quenches the fluorescence of the probe via photoinduced electron transfer to give a nonfluorescent species. Probes are selective for glutathione (GSH) in the red region and for cysteine/homocysteine (Cys/Hcy) in the green region. When they were treated with ...