Cysteine (Cys) is a bithiol that plays a vital role in many physiological processes. However, it is difficult to discriminate Cys from homocysteine (Hcy) and glutathione (GSH), due to their similar chemical structures and reactivity. Herein, we have developed a polymeric nanoprobe, nanoHFA, for ratiometric, highly selective, and sensitive detection of Cys based on 7-hydroxycoumarin-3-carboxylic acid (HC) and fluorescein isothiocyanate (FITC)-acrylate (FITC-A) group-functionalized lipopolymer DSPE-PEG. The probe nanoHFA showed a strong fluorescence emission peak centered at 450 nm attributed to HC and a weak fluorescence emission peak centered at 520 nm due to the photoinduced electron transfer (PET) process of FITC induced by acrylate group. In the presence of Cys, the fluorescence signal at 520 nm could be lit up and the ratio of F520nm/F450nm showed a good linear relationship in the range of 5–60 μM with a low detection limit of 0.37 μM. The probe also displayed excellent water solubility and high selectivity to Cys over other biothiols such as Hcy and GSH. Moreover, we further used probe nanoHFA to detect Cu2+ ions in the range of 100–550 nM with a detection limit of 77 nM. The nanoprobe was successfully applied for the quantitative detection of Cys in fetal bovine serum, and fluorescent strips were developed for facile and visual detection of Cys and Cu2+ ions.
This is a preview of subscription content, access via your institution.
Buy single article
Instant access to the full article PDF.
Price excludes VAT (USA)
Tax calculation will be finalised during checkout.
Wood ZA, Schröder E, Harris JR, Poole LB. Structure, mechanism and regulation of peroxiredoxins. Trends Biochem Sci. 2003;28(1):32–40.
Reddie KG, Carroll KS. Expanding the functional diversity of proteins through cysteine oxidation. Curr Opin Chem Biol. 2008;12(6):746–54.
Weerapana E, Wang C, Simon GM, Richter F, Khare S, Dillon MB, et al. Quantitative reactivity profiling predicts functional cysteines in proteomes. Nature. 2010;468(7325):790–5.
Li Q, Guo Y, Shao S. A BODIPY based fluorescent chemosensor for Cu (II) ions and homocysteine/cysteine. Sens Actuators B Chem. 2012;171:872–7.
Niu LY, Guan YS, Chen YZ, Wu LZ, Tung CH, Yang QZ. A turn-on fluorescent sensor for the discrimination of cystein from homocystein and glutathione. Chem Commun. 2013;49(13):1294–6.
Chwatko G, Bald E. Determination of cysteine in human plasma by high-performance liquid chromatography and ultraviolet detection after pre-column derivatization with 2-chloro-1-methylpyridinium iodide. Talanta. 2000;52(3):509–15.
Guan X, Hoffman B, Dwivedi C, Matthees DP. A simultaneous liquid chromatography/mass spectrometric assay of glutathione, cysteine, homocysteine and their disulfides in biological samples. J Pharm Biomed. 2003;31(2):251–61.
Tang X, Liu Y, Hou H, You T. Electrochemical determination of L-tryptophan, L-tyrosine and L-cysteine using electrospun carbon nanofibers modified electrode. Talanta. 2010;80(5):2182–6.
Chen X, Zhou Y, Peng X, Yoon J. Fluorescent and colorimetric probes for detection of thiols. Chem Soc Rev. 2010;39(6):2120–35.
Wang H, Zhou G, Gai H, Chen X. A fluorescein-based probe with high selectivity to cysteine over homocysteine and glutathione. Chem Commun. 2012;48(67):8341–3.
Xiang HJ, Tham HP, Nguyen MD, Phua SZF, Lim WQ, Liu JG, et al. An aza-BODIPY based near-infrared fluorescent probe for sensitive discrimination of cysteine/homocysteine and glutathione in living cells. Chem Commun. 2017;53(37):5220–3.
Yue Y, Huo F, Ning P, Zhang Y, Chao J, Meng X, et al. Dual-site fluorescent probe for visualizing the metabolism of Cys in living cells. J Am Chem Soc. 2017;139(8):3181–5.
Niu LY, Chen YZ, Zheng HR, Wu LZ, Tung CH, Yang QZ. Design strategies of fluorescent probes for selective detection among biothiols. Chem Soc Rev. 2015;44(17):6143–60.
Niu W, Guo L, Li Y, Shuang S, Dong C, Wong MS. Highly selective two-photon fluorescent probe for ratiometric sensing and imaging cysteine in mitochondria. Anal Chem. 2016;88(3):1908–14.
Yang X, Guo Y, Strongin RM. Conjugate addition/cyclization sequence enables selective and simultaneous fluorescence detection of cysteine and homocysteine. Angew Chem Int Ed. 2011;50(45):10690–3.
Kwon H, Lee K, Kim HJ. Coumarin–malonitrile conjugate as a fluorescence turn-on probe for biothiols and its cellular expression. Chem Commun. 2011;47(6):1773–5.
Liu Y, Lv X, Hou M, Shi Y, Guo W. Selective fluorescence detection of cysteine over homocysteine and glutathione based on a cysteine-triggered dual Michael addition/retro-aza-aldol cascade reaction. Anal Chem. 2015;87(22):11475–83.
Liu J, Sun YQ, Huo Y, Zhang H, Wang L, Zhang P, et al. Simultaneous fluorescence sensing of Cys and GSH from different emission channels. J Am Chem Soc. 2013;136(2):574–7.
Guo J, Yang S, Guo C, Zeng Q, Qing Z, Cao Z, et al. Molecular engineering of α-substituted acrylate ester template for efficient fluorescence probe of hydrogen polysulfides. Anal Chem. 2017;90(1):881–7.
Pang L, Zhou Y, Gao W, Zhang J, Song H, Wang X, et al. Curcumin-based fluorescent and colorimetric probe for detecting cysteine in living cells and zebrafish. Ind Eng Chem Res. 2017;56(27):7650–5.
Chen C, Liu W, Xu C, Liu W. A colorimetric and fluorescent probe for detecting intracellular biothiols. Biosens Bioelectron. 2016;85:46–52.
Rani BK, John SA. A novel pyrene based fluorescent probe for selective detection of cysteine in presence of other bio-thiols in living cells. Biosens Bioelectron. 2016;83:237–42.
Wang J, Li B, Zhao W, Zhang X, Luo X, Corkins ME, et al. Two-photon near infrared fluorescent turn-on probe toward cysteine and its imaging applications. ACS Sensor. 2016;1(7):882–7.
Fu ZH, Han X, Shao Y, Fang J, Zhang ZH, Wang YW, et al. Fluorescein-based chromogenic and ratiometric fluorescence probe for highly selective detection of cysteine and its application in bioimaging. Anal Chem. 2017;89(3):1937–44.
Gao B, Cui L, Pan Y, Zhang G, Zhou Y, Zhang C, et al. A highly selective ratiometric fluorescent probe for biothiol and imaging in live cells. RSC Adv. 2016;6(49):43028–33.
Lim CS, Masanta G, Kim HJ, Han JH, Kim HM, Cho BR. Ratiometric detection of mitochondrial thiols with a two-photon fluorescent probe. J Am Chem Soc. 2011;133(29):11132–5.
Cabral H, Matsumoto Y, Mizuno K, Chen Q, Murakami M, Kimura M, et al. Accumulation of sub-100 nm polymeric micelles in poorly permeable tumours depends on size. Nat Nanotechnol. 2011;6(12):815–23.
Bae Y, Fukushima S, Harada A, Kataoka K. Design of environment-sensitive supramolecular assemblies for intracellular drug delivery: polymeric micelles that are responsive to intracellular pH change. Angew Chem Int Edit. 2003;42(38):4640–3.
Lu L, Feng C, Xu J, Wang F, Yu H, Xu Z, et al. Hydrophobic-carbon-dot-based dual-emission micelle for ratiometric fluorescence biosensing and imaging of Cu2+ in liver cells. Biosens Bioelectron. 2017;92:101–8.
Pan L, Sun S, Zhang L, Jiang K, Lin H. Near-infrared emissive carbon dots for two-photon fluorescence bioimaging. Nanoscale. 2017;8(39):17350–6.
Song J, Li S, Zhao C, Lu Y, Zhao D, Sun J, et al. A superhydrophilic cement-coated mesh: an acid, alkali, and organic reagent-free material for oil/water separation. Nanoscale. 2018;10:1920–9.
Erdal NB, Hakkarainen M. Construction of bioactive and reinforced bioresorbable nanocomposites by reduced nano-graphene oxide carbon dots. Biomacromolecules. 2018;19:1074–81.
Domaille DW, Zeng L, Chang CJ. Visualizing ascorbate-triggered release of labile copper within living cells using a ratiometric fluorescent sensor. J Am Chem Soc. 2010;132(4):1194–5.
Siluvai GS, Mayfield M, Nilges MJ, DeBeer George S, Blackburn NJ. Anatomy of a red copper center: spectroscopic identification and reactivity of the copper centers of Bacillus subtilis Sco and its Cys-to-Ala variants. J Am Chem Soc. 2010;132(14):5215–26.
Wang YQ, Zhao T, He XW, Li WY, Zhang YK. A novel core-satellite CdTe/Silica/Au NCs hybrid sphere as dual-emission ratiometric fluorescent probe for Cu2+. Biosens Bioelectron. 2014;51:40–6.
Jo S, Kim D, Son SH, Kim Y, Lee TS. Conjugated poly (fluorene-quinoxaline) for fluorescence imaging and chemical detection of nerve agents with its paper-based strip. ACS Appl Mater Inter. 2014;6(2):1330–6.
You M, Lin M, Gong Y, Wang S, Li A, Ji L, et al. Household fluorescent lateral flow strip platform for sensitive and quantitative prognosis of heart failure using dual-color upconversion nanoparticles. ACS Nano. 2017;11(6):6261–70.
Yan X, Li H, Zheng W, Su X. Visual and fluorescent detection of tyrosinase activity by using a dual-emission ratiometric fluorescence probe. Anal Chem. 2015;87(17):8904–9.
This work was financially supported by the National Natural Science Foundation of China (21775046, 21675055, 21635003).
Conflict of interest
The authors declare that they have no conflict of interest.
Electronic supplementary material
About this article
Cite this article
Wang, F., Zhu, Y., Xu, J. et al. Highly selective and ratiometric fluorescent nanoprobe for the detection of cysteine and its application in test strips. Anal Bioanal Chem 410, 4875–4884 (2018). https://doi.org/10.1007/s00216-018-1128-1