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An ultrasensitive electrochemical DNA biosensor for human papillomavirus (HPV) detection was developed by electrochemical impedance spectroscopy and differential pulse voltammetry. A capture probe was immobilized on a glassy carbon electrode modified with graphene/Au nanorod/polythionine (G/Au NR/PT). Two auxiliary probes were designed and used to long-range self-assemble DNA nanostructure. The target DNA can connect DNA structure to the capture probe on the electrode surface. [Ru(phen) 3 ] 2+ was selected as a redox indicator for amplifying electrochemical signal significantly. Enhanced sensitivity was obtained through combining the excellent electric conductivity of G/Au NR/PT architecture and the long-range self-assembly DNA nanostructure with the multi-signal amplification. The DNA biosensor displayed excellent performance for HPV DNA detection over the range from 1.0×10 –13 to 1.0×10 –10 mol/L with a detection limit of 4.03×10 –14 mol/L. Furthermore, the proposed method can also be used for the detection of HPV DNA in human serum samples and provides a potential application of DNA detection in clinic research. [ABSTRACT FROM AUTHOR]

Well-dispersed graphene/gold nanorod (G/GNR) composites were synthesized by directly reducing a mixture of graphene and gold growth solution with sodium borohydride. The synthesized G/GNR composites were characterized by ultraviolet-visible spectroscopy and Fourier transform infrared spectroscopy, and the composites were observed by transmission electronic microscopy, which revealed that the GNRs were self-assembled onto the surface of graphene sheets. Glassy carbon electrodes were modified with G/GNR composites to construct a ractopamine electrochemical sensor. A sensitive, rapid, and simple electrochemical method was developed for the detection of ractopamine based on the strong enhancement effect of G/GNRs. The peak currents varied linearly with the concentration of ractopamine over the range of 1×10−9 to 2.7×10−6 molL−1, and the detection limit was 5.1×10−10 molL−1 (S/N=3). This method was applied to detect the content of ractopamine in swine urine samples, and the recovery was in the range of 99.2 to 107.3%. [ABSTRACT FROM AUTHOR]

Highlights • A convenient ECL biosensing method for selective detection of 5-hmC was developed. • The method is based on thiolated anchor probe DNA to immobilize 5-hmC-dsDNA. • The method can be used for detection of 5-hmC in genomic DNA. Abstract Herein, a novel and convenient electrogenerated chemiluminescence (ECL) biosensing method for the quantitative detection of 5-hydroxymethylcytosine (5-hmC) in genomic DNA was developed by employing thiolated anchor probe DNA to immobilize 5-hydroxymethylcytosine double-stranded DNA (5-hmC-dsDNA) and a ruthenium(II) complex-tagged reporter probe as an ECL probe. Briefly, the 5-hmC-dsDNA was firstly oxidized to 5-formylcytosine dsDNA (5-fC-dsDNA) by KRuO 4 in high yields. Subsequently, the thiolated anchor probe DNA is employed to help immobilize the 5-fC-dsDNA onto the gold electrode surface via the Au-S bond. An ECL probe DNA, labeled with a signal transducer − Ru(bpy) 2 (cbpy)NHS, hybridized with the anchor probe DNA on the electrode, resulting in an intensive ECL emission. The obtained ECL intensity was linearly proportional to the content of 5-hmC in the range from 0.0090% to 0.5761%, with the limit of detection calculated to be 0.0036% (S/N = 3). Finally, we applied the developed 5-hmC detection strategy in various mice tissues. It demonstrates that the content of 5-hmC measured by this method in different mice tissues vary significantly: 5-hmC is highest in brain and lowest in spleen. The success in assaying the mice tissues samples signifies its great promise in a broad range of practical applications for the quantitative detection of 5-hmC in genomic DNA. [ABSTRACT FROM AUTHOR]

Abstract 5-Hydroxymethylcytosine (5-hmC), an oxidation product of 5-mC (5-methylcytosine), is presented in DNA of most mammalian cells and play an important role in the alteration of cancer-related genes. Herein, a sensitive electrogenerated chemiluminescence (ECL) biosensing method for the determination of 5-hmC in DNA (5-hmC DNA) was established on the basis of chemical modification and nanomaterial amplification. First, electrochemically reduced molybdenum disulfide-poly(acrylic acid) (rMoS 2 -PAA) nanosheets were used to modify glassy carbon electrode (GCE) to form an ECL biosensing electrode (rMoS 2 -PAA/GCE) which has large accessible surface area to immobilize more DNA. Then, a capture probe with amino group was hybridized with the target 5-hmC DNA and immobilized on the surface of rMoS 2 -PAA/GCE via amido bond. When cysteamine was introduced, the M.HhaI methyltransferase (M.HhaI) was used as specific recognition element to replace the hydroxyl group of 5-hmC by thiol and generated the amine-derivated DNA. Finally, surface chemically activated Ru(bpy) 3 2+-doped silica (Ru@SiO 2) nanoparticles, carriers of ECL reagents, were employed as signal amplification unit which covalently bonded to the amine-derivated DNA resulting in an increased ECL intensity. The increased ECL intensity was linearity to the 5-hmC DNA concentration in a range from 5.0 × 10−14 M to 1.0 × 10−11 M, with a lower detection limit of 1.2 × 10−14 M. Besides, the proposed method also displayed a good selectivity to 5-hmC in the presence of 5-C and 5-mC. Moreover, the developed biosensing method was successfully employed to monitor human urine sample. Graphical abstract fx1 Highlights • An ECL biosensing method for assay of the DNA hydroxymethylation was developed. • The method is based on M. HhaI for chemical modification 5-hmC-DNA. • The amplification of MoS 2 -PAA and Ru@SiO 2 lead to a favorable sensitivity. • The method achieved a lower DL of 1.2 × 10−14 M for detection of 5-hmC-DNA. [ABSTRACT FROM AUTHOR]

MicroRNA-21 (miRNA-21) is a promising diagnostic biomarker for breast cancer screening and disease progression. A sensitive and selective strategy for the quantitative determination of miRNA-21 is of great significance in the early diagnosis of cancers. Herein, a novel electrogenerated chemiluminescence (ECL) biosensor was designed for the detection of miRNA-21 with dual signal amplification based on isothermal strand-displacement polymerase reaction (ISDPR) and bridge DNA-gold nanoparticles (AuNPs) nanocomposites. The ECL biosensor was fabricated by self-assembling a thiolated capture probe (SH-CP) on the surface of a gold electrode. The target miRNA-21 initiated the phi29 DNA polymerase-mediated ISDPR, which could generate large numbers of single-stranded DNA (assistant DNA) with accurate and comprehensive nucleotide sequences. The assistant DNA was captured by the SH-CP self-assembled on the Au electrode and further hybridized with bridge DNA-AuNPs nanocomposites, more biotins can be captured on the electrode surface. Afterward, a streptavidin-modified Ru (bpy) 3 2+ complex (SA-Ru) was bound to the bridge DNA-AuNPs nanocomposites via a specific interaction between biotin and streptavidin to produce a strong ECL signal. The ECL intensity was logarithmically proportion to the concentration of target miRNA-21 over a range from 0.01 fM to 10,000 fM with a detection limit of 3.2 aM. The proposed ECL biosensor was successfully applied to detect miRNA-21 in total RNA samples extracted from human breast cancer cells, and it showed great potential for early cancer diagnosis based on miRNA as a biomarker. • An ultrasensitive ECL biosensor for the detection of microRNA was developed. • The target triggered the recycling reaction and bridge DNA-AuNPs were utilized to realize efficient signal amplification. • The ECL biosensor can be used for detection of miRNA-21 in total RNA samples extracted from human breast cancer cells. [ABSTRACT FROM AUTHOR]