A novel optical DNA biosensor for detection of trace concentration of Methylene blue using Gold nano-particles and Guanine rich single strand DNA

Document Type: Reasearch Paper


Faculty of Chemistry, Kharazmi University, Tehran, Iran


The glass surface modification with 3-(mercaptopropyl) trimethoxysilane (MPTS), gold nano-particles (GN) and guanine rich single strand DNA (ss-DNA) was utilized as a novel and efficient platform for sensing trace concentration of methylene blue (MB) by an inexpensive spectrophotometric method. Methylene Blue (MB) can interact with the guanine base of single strand DNA and absorbed onto glass surface and its absorbance can be determined at λmax of MB. Increase in methylene blue absorbance was linear with an increase in MB concentration and can be viewed by the naked eye and detected by spectrophotometric method. The linear range of the developed biosensor for determination of MB was from 10 to 100 nmol L-1 with a detection limit of 4 nmol L-1. The fabricated spectrophotometric MB biosensor possessed excellent selectivity and good sensitivity.


Main Subjects

[1] Wheeler M. M., Wade J. S. H., (1982), Intraoperative identification of parathyroid glands: Appraisal of methylene blue staining. Am. J. Surg. 143: 713-716.

[2] Milani A., Ciammella A. M., Degen C., Siciliano M., Rossi L., (1992), Ascites dynamics in cirrhosis. Proposal and validation of a methylene blue dilution test. J. Hepatol. 16: 369-375.

[3] Noodt B. B., Rodal G. H., Wainwright M., Peng Q., Horobin R., Nesland J. M., Berg K., (1998),  Apoptosis induction by different pathways with methylene blue derivative and light from mitochondrial sites in V9 cells. Int. J. Cancer. 75: 941-948.

[4] Turnipseed S. B., Roybal J. E., Plakas S. M., Pfenning A. P., Hurlbut J. A., Long A. R., (1997), Determination of methylene blue in channel catfish tissue by liquis chromatography with visible detection. J. AOAC. Int. 80: 31-35.

[5] Borwitzky H., Haefeli W. E., Burhenne J., (2005), Analysis of methylene blue in human urine by capillary electrophoresis. J. Chromatogr. B. 826: 244-251.

[6] Kolekar Y. M., Pawar S. P., Gawai K. R., Lokhande P. D., Shouche Y. S., Kodam K. M., (2008), Decolorization and degradation of Disperse Blue 79 and Acid Orange 10, by Bacillus fusiformis KMK5 isolated from the textile dye contaminated soil. Bioresour. Technol. 99: 8999-9003.

[7] Muthuraman G., Teng T. T., (2009), Extraction and recovery of rhodamine B, methyl violet and methylene blue from industrial wastewater using D2EHPA as an extractant. J. Ind. Eng. Chem. 15: 841-846.  

[8] Burhenne J., Riedel K. D., Rengelshausen J., Meissner P., Müller Q., Mikus G., Haefeli W. E., Walter-Sack I., (2008), Quantification of cationic anti-malaria agent methylene blue in different human biological matrices using cation exchange chromatography coupled to tandem mass spectrometry. J. Chromatogr. B. 863: 273-282.

[9] Rukmini N., Kavitha V. S., (1978), Reductimetric titration of methylene blue, thionine, nitroso-R-salt, 1-nitroso-2-naphthol and 2-nitroso-1-naphthol. J. Indian. Chem. Soc. 55: 660-661.

[10] Pamfilov A. V., Ya S., Mazurkevitch X., Pakhomova E. P., (1968), Mechanism of reduction of methylene blue and thionine. Ukr. Khim. Zh. 34: 276-280.

[11] Selig, W., (1981), Potentiometric titration of organic cations precipitated by tetraphenylborate. Fresenius Z. Anal. Chem. 308: 21-23.

[12] Orwitzky H., Haefelh W. F., Burhenne J., (2005), Analysis of methylene blue in human urine by capillary electrophoresis. J. Chromatogr. B. 826: 244-251.

[13] Xu J., Dai L., Wu B., Ding T., Zhu J. J., Lin H., (2009), Determination of methylene blue residues in aquatic products by liquid chromatography-tandem mass spectrometry. J. Sep. Sci. 32: 4193-4199.

[14] Zhao Y., Chen X., Wang C. Y., (1998), A mehylene blue-selective membrane electrode using methylene blue-phosphotungstate as electroactive material and its pharmaceutical applications. Croat. Chim. Acta. 71: 757-764.

[15] Lachheb H., Puzenat E., Houas A., Ksibi M., Elaloubi E., Gyillard C., Herrmann J. M., (2002), Photocatalytic degradation of various types of dyes (Alizarin S, Crocein Orange G, Methyl Red, Congo Red, Methylene Blue) in water by UV-irradiated titania. Appl. Catal. B. 39: 75-90.

[16] Chitikela S., Dentel S. K., Aleen H., (1995), Modified method for the analysis of anionic surfactants as Methylene Blue active substances. Analyst. 120: 2001-2004.

[17] Yang F., Xia S., Liu Z., Chen J., Lin Y., Qiu B., Chen G., (2011), Analysis of methylene blue and its metabolites in blood by capillary electrophoresis/electrospray ionization mass spectrometry. Electrophoresis. 32: 659-664.

[18] Chiorcea Paquim A. M., Diculescu V. C., Oretskaya T. S., Oliveira Brett A. M., (2004), AFM and electroanalytical studies of synthetic oligonucleotide hybridization. Biosens. Bioelectron. 20: 933-944.

[19] Kerman K., Ozhan D., Kara P., Meric B., Gooding J. J., Ozsoz M., (2002), Voltammetric determination of DNA hybridization using methylene blue and self-assembled alkanethiol monolayer on gold electrodes. Anal. Chim. Acta. 462: 39-47.

[20] Mashhadizadeh M. H., Talemi R. P., (2013), A new methodology for electrostatic immobilization of a non-labeled single strand DNA onto a self-assembled diazonium modified gold electrode and detection of its hybridization by differential pulse voltammetry. Talanta. 103: 344-348.

[21] Sanchez-martinez M. L., Aguilar-Caballos A., Gomez-hens A., (2009), Homogeneous immunoassay for soy protein determination in food samples using gold nanoparticles as labels and light scattering detection. Anal. Chim. Acta. 63: 58-62.

[22] De M., Ghosh P. S., Rotello V. M., (2008), Applications of nanoparticles in biology. Adv. Mater. 20: 4225-4241.

[23] Mannelli I., Marco M. P., (2010), Recent advances in analytical and bioanalysis application of noble metal nanorods. Anal. Bioanal. Chem. 398: 2451-2469.

[24] Erdem A., Kerman K., Meric B., Akarca U. S., Ozsoz M., (2000), Novel hybridization indicator methylene blue for the electrochemical detection of short DNA sequences related to the hepatitis B virus. Anal. Chim. Acta. 422: 139-149.

[25] Meric B., Kerman K., Ozkan D., Kara P., Erensoy S., Akarca U. S., Mascini M., Ozsoz M., (2002), Electrochemical DNA biosensor for the detection of TT and Hepatitis B virus from PCR amplified real samples by using methylene blue. Talanta.  56: 837-846.

[26] The H. F., Gong H., Dong X. D., Zeng X., Lai Kuan Tan A., Yang X., Tan S. N., (2005), Electrochemical biosensing of DNA with capture probe covalently immobilized onto glassy carbon surface. Anal. Chim. Acta. 551: 23-29.

[27] Boon E. M., Ceres D. M., Drummond T. G., Hill M. G., Barton J. K., (2000), Mutation detection by electrocatalysis at DNA-modified electrodes. Nat. Biotech. 18: 1096-1100.

[28] Kelley S. O., Barton J. K., Jackson N. M., Hill M. G., (1997), Electrochemistry of methylene blue bound to a DNA-modified electrode. Bioconj. Chem. 8: 31-37.

[29] Wang J., Wang F., Dong S., (2009), Methylene blue as an indicator for sensitive electrochemical detection of adenosine based on aptamer switch. J. Electroanal. Chem.  626: 1-5.

[30] Arzum E., Kagan K., Burcu M., Mehmet O., (2001), Methylene blue as a novel electrochemical hybridization indicator. Electroanalysis. 13: 219-223.

[31] Christine B., Per S., Jan B., Gillis J., (1999), A feasibility study of a capacitive biosensor for direct detection of DNA hybridization. Electroanalysis. 11: 156-160.

[32] Mashhadizadeh M. H., Talemi R. P., (2014), A novel optical DNA biosensor for detection of trace amounts of mercuric ions using gold nanoparticles introduced onto modified glass surface. Spectrochim. Acta A. 132: 403-409.

[33] Mashhadizadeh M. H., Talemi R. P., (2011), Used gold nano-particles as an on/off switch for response of a potentiometric sensor to Al (III) or Cu (II) metal ions. Anal. Chim. Acta. 692: 109-115.

[34] Fujiwara K., Kasaya H., Ogawa N., (2009), Gold nanoparticle monolayer formation on a chemically modified Glass Surface. Anal. Sci. 25: 241-248.

[35] Zhu L., Zhao R., Wang K., Xiang H., Shang Z., Sun W., (2008), Electrochemical behaviors of methylene blue on DNA modified electrode and Its application to the detection of PCR product from NOS sequence. Sensors. 8: 5649-5660.

[36] Nafisi S., Saboury A. A., Keramat N., Neault J. F., Tajmir-Riahi H. A., (2007), Stability and structural features of DNA intercalation with ethidium bromide, acridine orange and methylene blue. J. Mol. Struc. 827: 35-43.

[37] Belaz-Davld N., Decosterd L. A. , Appenzeller M., Ruetsch Y., Rhiolro H.,  Buchn T., Biollaz J., (1997), Spectrophotometric determination of methylene blue in biological fluids after ion-pair extraction and evidence of its adsorption on plastic polymers. European J. Pharm. Sci. 5: 335-345.

[38] Wen M. L., Zhao Y. B., Chen X., Wang C. Y., (1999), Potentiometric sensor for methylene blue based on methylene blue–silicotung state ion association and its pharmaceutical applications. J. Pharm. Biomed. Analysis. 18: 957–961.

[39] Rizwan Khan M., Ali Khan M., Alothman Z. A., Alsohaimi A. H., Naushad M., Al-Shaalan N. H., (2014), Quantitative determination of methylene blue in environmental samples by solid-phase extraction and ultra-performance liquid chromatographyt and mass spectrometry: A green approach. RSC Adv. 4: 34037-34044.