Abstract
An enzyme-free aptameric nanosensor is presented for apoptosis assay. The method exploits the peroxidase-mimicking property of silver/platinum alloy nanoclusters (Ag/Pt NCs) and uses a Cyt c binding ssDNA aptamer. An extra-strand polycytosine (C14) aptamer was designed as a template for synthesis of the Ag/Pt NCs. If cell lysate or purified Cyt c is placed in a polystyrene microplate, Cyt c will bind to the surface of the wells of a microtiterplate. On addition of Apt@Ag/PtNCs, it will associate with Cyt c and then catalytically oxidize colorless tetramethylbenzidine (TMB) in the presence of H2O2 to give a blue colored oxidation product (TMBox) due to the peroxidase-mimicking property of the Ag/Pt NCs. Under optimal conditions, the absorbance of TMB at 660 nm is linearly enhanced as the concentration of Cyt c increases from 50.0 fM to 500 nM, and the detection limit is ~10 pM. The assay is simple, sensitive and cost effective in that it is enzyme-free, antibody-free and label-free.
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Nur-E-Kamal A, Gross SR, Pan Z, Balklava Z, Ma J, Liu LF (2004) Nuclear translocation of cytochrome c during apoptosis. J Biol Chem 279:24911–24914
Wong RS (2011) Apoptosis in cancer: from pathogenesis to treatment. J Exp Clin Cancer Res 30:87
Lowe SW, Lin AW (2000) Apoptosis in cancer. Carcinogenesis 21:485–495
Wang C, Youle RJ (2009) The role of mitochondria in apoptosis. Annu Rev Genet 43:95–118
Radhakrishnan J, Origenes R, Littlejohn G, Nikolich S, Choi E, Smite S, Lamoureux L, Baetiong A, Shah M, Gazmuri RJ (2017) Plasma cytochrome c detection using a highly sensitive electrochemiluminescence enzyme-linked immunosorbent assay. Biomark Insights 12:1177271917746972
Gao W, Pu Y, Luo KQ, Chang DC (2001) Temporal relationship between cytochrome c release and mitochondrial swelling during UV-induced apoptosis in living HeLa cells. J Cell Sci 114:2855–2862
Brown GC, Borutaite V (2008) Regulation of apoptosis by the redox state of cytochrome c. Biochimica et Biophysica Acta (BBA)-Bioenergetics 1777:877–881
Jiang X, Wang X (2004) Cytochrome C-mediated apoptosis. Annu Rev Biochem 9:73
Chandra D, Liu JW, Tang DG (2002) Early mitochondrial activation and cytochrome c up-regulation during apoptosis. J Biol Chem 277:50842–50854
Liu K, Shu D, Song N, Gai Z, Yuan Y, Li J, Li M, Guo S, Peng J, Hong H (2012) The role of cytochrome c on apoptosis induced by Anagrapha falcifera multiple nuclear polyhedrosis virus in insect Spodoptera litura cells. PLoS One 7:e40877
Torkzadeh-Mahani M, Ataei F, Nikkhah M, Hosseinkhani S (2012) Design and development of a whole-cell luminescent biosensor for detection of early-stage of apoptosis. Biosens Bioelectron 38:362–368
Noori AR, Hosseini ES, Nikkhah M, Hosseinkhani S (2018) Apoptosome formation upon overexpression of native and truncated Apaf-1 in cell-free and cell-based systems. Arch Biochem Biophys 642:46–51
Akbari-Birgani S, Hosseinkhani S, Mollamohamadi S, Baharvand H (2014) Delay in apoptosome formation attenuates apoptosis in mouse embryonic stem cell differentiation. J Biol Chem 289:16905–16913
Bin N, Li W, Yin X, Huang X, Cai Q (2016) Electrochemiluminescence aptasensor of TiO2/CdS: Mn hybrids for ultrasensitive detection of cytochrome c. Talanta 160:570–576
Ghayyem S, Faridbod F (2018) A fluorescent aptamer/carbon dots based assay for cytochrome c protein detection as a biomarker of cell apoptosis. Methods and applications in fluorescence 7:015005
Amin RM, Elfeky SA, Verwanger T, Krammer B (2017) Fluorescence-based CdTe nanosensor for sensitive detection of cytochrome C. Biosens Bioelectron 98:415–420
Chen TT, Tian X, Liu CL, Ge J, Chu X, Li Y (2015) Fluorescence activation imaging of cytochrome c released from mitochondria using aptameric nanosensor. J Am Chem Soc 137:982–989
Shamsipur M, Molaabasi F, Hosseinkhani S, Rahmati F (2016) Detection of early stage apoptotic cells based on label-free cytochrome c assay using bioconjugated metal nanoclusters as fluorescent probes. Anal Chem 88:2188–2197
Shamsipur M, Pashabadi A, Molaabasi F, Hosseinkhani S (2017) Impedimetric monitoring of apoptosis using cytochrome-aptamer bioconjugated silver nanocluster. Biosens Bioelectron 90:195–202
Chattoraj S, Amin MA, Bhattacharyya K (2016) Cytochrome c-capped fluorescent gold Nanoclusters: imaging of live cells and delivery of cytochrome c. ChemPhysChem 17:2088–2095
Pur MR, Hosseini M, Faridbod F, Ganjali MR, Hosseinkhani S (2018) Early detection of cell apoptosis by a cytochrome C label-free electrochemiluminescence aptasensor. Sensors Actuators B Chem 257:87–95
Pandiaraj M, Madasamy T, Gollavilli PN, Balamurugan M, Kotamraju S, Rao VK, Bhargava K, Karunakaran C (2013) Nanomaterial-based electrochemical biosensors for cytochrome c using cytochrome c reductase. Bioelectrochemistry 91:1–7
Santra S, Kaittanis C, Perez JM (2010) Cytochrome C encapsulating theranostic nanoparticles: a novel bifunctional system for targeted delivery of therapeutic membrane-impermeable proteins to tumors and imaging of cancer therapy. Mol Pharm 7:1209–1222
Hosseini M, Mohammadi S, Borghei YS, Ganjali MR Detection of p53 gene mutation (single-base mismatch) using a fluorescent silver nanoclusters. J Fluoresc 27:1443–1448
Borghei YS, Hosseini M, Ganjali MR, Ju H (2018) Colorimetric and energy transfer based fluorometric turn-on method for determination of microRNA using silver nanoclusters and gold nanoparticles. Microchim Acta 185:286
Borghei YS, Hosseini M, Ganjali MR (2018) Oxidase-like catalytic activity of Cys-AuNCs upon visible light irradiation and its application for visual miRNA detection. Sensors Actuators B Chem 273:1618–1626
Borghei YS, Hosseini M, Ganjali MR (2017) Detection of large deletion in human BRCA1 gene in human breast carcinoma MCF-7 cells by using DNA-silver Nanoclusters. Methods and applications in fluorescence 6:015001
Darabdhara G, Boruah PK, Das MR (2019) Colorimetric determination of glucose in solution and via the use of a paper strip by exploiting the peroxidase and oxidase mimicking activity of bimetallic cu-Pd nanoparticles deposited on reduced graphene oxide, graphitic carbon nitride, or MoS 2 nanosheets. Microchim Acta 186:13
Wang YW, Liu Q, Wang L, Tang S, Yang HH, Song H (2019) A colorimetric mercury (II) assay based on the hg (II)-stimulated peroxidase mimicking activity of a nanocomposite prepared from graphitic carbon nitride and gold nanoparticles. Microchim Acta 186:7
Song W, Yin W, Zhang Z, He P, Yang X, Zhang X (2019) A DNA functionalized porphyrinic metal-organic framework as a peroxidase mimicking catalyst for amperometric determination of the activity of T4 polynucleotide kinase. Microchim Acta 186:149
Zheng C, Zheng AX, Liu B, Zhang XL, He Y, Li J, Yang HH, Chen G (2014) One-pot synthesized DNA-templated Ag/Pt bimetallic nanoclusters as peroxidase mimics for colorimetric detection of thrombin. Chem Commun 50:13103–13106
Borghei YS, Hosseini M, Ganjali MR (2018) Visual detection of miRNA using peroxidase-like catalytic activity of DNA-CuNCs and methylene blue as indicator. Clin Chim Acta 483:119–125
Borghei YS, Hosseini M, Ganjali MR (2017) Fluorometric determination of microRNA via FRET between silver nanoclusters and CdTe quantum dots. Microchim Acta 184:4713–4721
Nasir M, Nawaz MH, Latif U, Yaqub M, Hayat A, Rahim A (2017) An overview on enzyme-mimicking nanomaterials for use in electrochemical and optical assays. Microchim Acta 184:323–342
Zuker M (2003) Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res 31:3406–3415
Sha H, Zhang Y, Wang Y, Ke H, Xiong X, Xue H, Jia N (2019) Electroluminescent aptasensor based on RuSiO2 nanoparticles for detection cytochrome c using ferrocene as quenching probe. Biosens Bioelectron 132:203–209
Acknowledgements
Financial support of this work was provided by National Institute for Medical Research Development (NIMAD, Grant No: 957982). The authors are grateful to the Iran National Elites Foundation for support of Yasaman-Sadat Borghei as a post-doc fellow in this work.
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Borghei, YS., Hosseinkhani, S. Aptamer-based colorimetric determination of early-stage apoptotic cells via the release of cytochrome c from mitochondria and by exploiting silver/platinum alloy nanoclusters as a peroxidase mimic. Microchim Acta 186, 845 (2019). https://doi.org/10.1007/s00604-019-3977-5
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DOI: https://doi.org/10.1007/s00604-019-3977-5