An electrochemical biosensor for the amplification of thrombin activity by perylene-mediated photoinitiated polymerization.

BACKGROUND: Thrombin, a coagulation system protease, is a key enzyme involved in the coagulation cascade and has been developed as a marker for coagulation disorders. However, the methods developed in recent years have the disadvantages of complex operation, long reaction time, low specificity and sensitivity. Meanwhile, thrombin is at a lower level in the pre-disease period. Therefore, to accurately diagnose the disease, it is necessary to develop a fast, simple, highly sensitive and specific method using signal amplification technology. RESULTS: We designed an electrochemical biosensor based on photocatalytic atom transfer radical polymerization (photo-ATRP) signal amplification for the detection of thrombin. Sulfhydryl substrate peptides (without carboxyl groups) are self-assembled to the gold electrode surface via Au-S bond and serve as thrombin recognition probes. The substrate peptide is cleaved in the presence of thrombin to generate -COOH, which can form a carboxylate-Zr(IV)-carboxylate complex via Zr(IV) and initiator (α-bromophenylacetic acid, BPAA). Subsequently, an electrochemical biosensor was prepared by introducing polymer chains with electrochemical signaling molecules (ferrocene, Fc) onto the electrode surface by photocatalytic (perylene, Py) mediated ATRP using ferrocenylmethyl methacrylate (FMMA) as a monomer. The concentration of thrombin was evaluated by the voltammetric signal generated by square wave voltammetry (SWV), and the result showed that the biosensor was linear between 1.0 ng/mL âˆ¼ 10 fg/mL, with a lower detection limit of 4.0 fg/mL (∼0.1 fM). Moreover, it was shown to be highly selective for thrombin activity in complex serum samples and for thrombin inhibition screening. SIGNIFICANCE: The biosensor is an environmentally friendly and economically efficient strategy while maintaining the advantages of high sensitivity, anti-interference, good stability and simplicity of operation, which has great potential for application in the analysis of complex samples.

Ultrasensitive detection of miRNA-21 by click chemistry and fluorescein-mediated photo-ATRP signal amplification.

The development of a convenient and efficient assay using miRNA-21 as a lung cancer marker is of great importance for the early prevention of cancer. Herein, an electrochemical biosensor for the detection of miRNA-21 was successfully fabricated under blue light excitation using click chemistry and photocatalytic atom transfer radical polymerization (photo-ATRP). By using hairpin DNA as a recognition probe, the electrochemical sensor deposits numerous electroactive monomers (ferrocenylmethyl methacrylate) on the electrode surface under the reaction of photocatalyst (fluorescein) and pentamethyldiethylenetriamine, thereby achieving signal amplification. This biosensor is sensitive, precise and selective for miRNA-21, and is highly specific for RNAs with different base mismatches. Under optimal conditions, the biosensor showed a linear relationship in the range of 10 fM ∼1 nM (R2 = 0.995), with a detection limit of 1.35 fM. Furthermore, the biosensor exhibits anti-interference performance when analyzing RNAs in serum samples. The biosensor is based on green chemistry and has the advantages of low cost, specificity and anti-interference ability, providing economic benefits while achieving detection objectives, which makes it highly promising for the analysis of complex samples.

Solvent-directed multiple correspondence fluorescent probe for highly selective and sensitive detection of Cu2+ and Mg2.

A solvent-directed, new Schiff base multiple correspondence fluorescent probe, (E)-2-(2-hydroxybenzylidene) hydrazine-1-carboxamid (L), was synthesized for selective sensing of Cu2+ and Mg2+ ions. L showed excellent selectivity and high sensitivity toward Cu2+ in "turn off" mode with a detection limit of 40.5 nM in 10 mM, pH = 7.0 PBS buffer. Contrary to that, when acetonitrile was used as the solvent, L exhibited highly selective and sensitive fluorescence sensing ability for Mg2+ in "turn on" mode with a detection limit of 9.5 nM. L can coordinate to Cu2+ and Mg2+ in a 1:1 molar ratio, respectively, evidenced by Job's plot analysis. Their binding modes were investigated by NMR, IR and XPS spectroscopies. Moreover, the satisfied results were obtained when L was used to detect Cu2+ and Mg2+ in real water samples.

Cyanocobalamin (VB12) Bionic Enzyme-Assisted Photocatalytic Chain-Growth Polymerization for Detection of Lung Cancer Biomarkers.

Cobalt-mediated radical polymerization is noted for its great level of control over the polymerization of acrylic and vinyl esters monomers, even at high molar mass. Vitamin B12, a natural bionic enzyme cobalt complex, involves the conversion of organic halides to olefins through chain-growth polymerization. In this work, the notion of R-Co(III) free radical persistent free radical effect and vitamin B12 circulation were first reported for the perception of ultralow abundance of microRNA-21, a lung cancer biomarker. Indeed, most Co-containing catalytic reactions can occur under mild conditions due to their minimal bond dissociation of the C-Co bond, with blue light irradiation. Based on the intrinsic stability of the vitamin B12 framework and recycling of the catalyst, it is evident that this natural catalytic scheme has potential applications in medicinal chemistry and biomaterials. In addition, this strategy, combined with highly specific recognition probes and vitamin B12 circulation-mediated chain-growth polymerization, has a detection limit as low as 910 aM. Furthermore, it is sensitive for sensing in serum samples containing biomarkers and shows great potential for RNA selection and amplification sensing in clinical samples.

Rose bengal-mediated photoinduced atom transfer radical polymerization for high sensitivity detection of target DNA.

Convenient and sensitive detection of biomolecules is of utmost importance in the field of early disease screening. In this study, a Rose Bengal-Mediated photoinduced atom transfer radical polymerization (photoATRP) method was used to achieve highly sensitive detection of target DNA (tDNA). The tDNA was specifically recognized using PNA with terminal modified sulfhydryl groups, and the initiator α-bromophenylacetic acid (BPAA) was attached to the electrode surface via a phosphate-Zr4+-carboxylate acid structure. Under the excitation of blue light, rose bengal (RB) acts as a photocatalyst, ß-nicotinamide adenine dinucleotide (NADH) as an electron donor, and ferrocenylmethyl methacrylate (FMMA) as a monomer to activate the photoATRP reaction and generate a large number of electroactive polymer chains on the electrode surface. Under optimal conditions, the method can be used for the quantitative analysis of tDNA in the concentration range of 1-105 fM (R2 = 0.994) with a limit of detection (LOD) of 0.115 fM. This metal-free mediated photoATRP biosensor, with low cost and environmental friendliness, has great potential in the field of highly sensitive biomolecule detection.

A novel colorimetric detection of glutathione based on stable free radical TEMPO oxidation of 3,3',5,5'-tetramethylbenzizine (TMB) via Copper(II) acetylacetonate catalysis.

In this work, a new colorimetric method for the determination of Glutathione (GSH) on the basis of stable free radical 2,2,6,6 - tetramethylpiperidine - 1 - oxyl (TEMPO) oxidation of 3,3',5,5'-tetramethylbenzizine (TMB) via copper(II) acetylacetonate (Cu(acac)2) catalysis was proposed. TEMPO was catalyzed by Cu(acac)2 to produce TEMPO+, then TEMPO+ oxidized TMB to produce oxidized TMB (ox - TMB). The resulting ox - TMB showed blue and possessed a distinct absorption peak about 650 nm. Whereas, GSH prohibited the generation of ox - TMB through inhibiting TMB oxidation. As compared to the case that GSH was absent, significantly enhanced absorption was determined, and was proportional to GSH amount. On this basis, a qualitative and quantitative detection method of GSH with the naked eye and the microplate reader was achieved. The developed TEMPO - based method achieved GSH biosensing with improved sensitivity in a good specificity - manner. The limit of detection (LOD) was 90 µM via naked eye, and the microplate reader was 4.71 µM. And the stable free radical TEMPO possessed higher stability and lower toxicity than traditional oxidant of H2O2. Moreover, this TEMPO - based method achieved good results in the detection of GSH in human serums.

Research Progress in Fluorescent Probes for Arsenic Species.

Arsenic is a toxic non-metallic element that is widely found in nature. In addition, arsenic and arsenic compounds are included in the list of Group I carcinogens and toxic water pollutants. Therefore, rapid and efficient methods for detecting arsenic are necessary. In the past decade, a variety of small molecule fluorescent probes have been developed, which has been widely recognized for their rapidness, efficiency, convenience and sensitivity. With the development of new nanomaterials (AuNPs, CDs and QDs), organic molecules and biomolecules, the conventional detection of arsenic species based on fluorescence spectroscopy is gradually transforming from the laboratory to the portable kit. Therefore, in view of the current research status, this review introduces the research progress of both traditional and newly developed fluorescence spectrometry based on novel materials for arsenic detection, and discusses the potential of this technology in the rapid screening and field testing of water samples contaminated with arsenic. The review also discusses the problems that still exist in this field, as well as the expectations.

Sensitive electrochemiluminescence analysis of lung cancer marker miRNA-21 based on RAFT signal amplification.

An electrochemiluminescence approach based on surface-initiated reversible addition-fragmentation chain transfer (SI-RAFT) was developed for miRNA-21 detection for the first time. The SI-RAFT polymerization generates polymer chains with functional groups that are used to recruit luminol, enabling strong ECL signal output with low concentrations of miRNA-21, and greatly improving the detection sensitivity.

A highly selective and sensitive Zn2+ fluorescent sensor based on zinc finger-like peptide and its application in cell imaging.

Developing new chemosensors for detection of Zn2+ has attracted great attentions because of the important roles of Zn2+ in biological systems, and it will produce toxic effects with an excessive intake of zinc ion. Metalloproteins are often used as an effective template for the design and development of peptide-based fluorescent sensors. In this study, we designed a new and simple ratiometric fluorescent sensor for Zn2+, which was based on a zinc finger-like peptide and labeled with a dansyl group, i.e., Dansyl-His-Gln-Arg-Thr-His-Trp-NH2 (D-P6), by using solid phase peptide synthesis (SPPS). The dimeric peptide has a high affinity for Zn2+ overothermetalions, as indicated by spectroscopic studies, as well as molecular modeling. Remarkably, the sensor exhibited a highly selective and sensitive ratiometric fluorescent response to Zn2+ by fluorescent resonance energy transfer effect between tryptophan residue and fluorophore dansyl group, with a very low detection limit of 33 nM in aqueous solution. Furthermore, the sensor displayed a very low biotoxicity, which allows successful detection of Zn2+ in living HeLa cells. We believe that the new sensor may have potential applications in biological science.

A Highly Selective and Sensitive Peptide-Based Fluorescent Ratio Sensor for Ag.

A fluorescence ratio sensor based on dansyl-peptide, Dansyl-Glu-Cys-Glu-Glu-Trp-NH2 (D-P5), was efficiently synthesized by Fmoc solid phase peptide synthesis. The sensor exhibits high selectivity and sensitivity for Ag+ over 16 metal ions in 100 mM sodium perchlorate and 50 mM 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid buffer solution by fluorescence resonance energy transfer. The 1:1 binding stoichiometry of the sensor and Ag+ is measured by fluorescence ratio response and the job's plot. The dissociation constant of the sensor with Ag+ was calculated to be 6.4 × 10-9 M, which indicates that the sensor has an effective binding affinity for Ag+. In addition, the limit of detection of the sensor for Ag+ was determined to be 80 nM, which also indicates that the sensor has a high sensitivity to Ag+. Result showed that the sensor is an excellent Ag+ sensor under neutral condition. Furthermore, this sensor displays good practicality for Ag+ detection in river water samples without performing tedious sample pretreatment, as well as for silver chloride detection.

Ultrasensitive mercury ion and biothiol detection based on Dansyl-His-Pro-Gly-Asp-NH2 fluorescent sensor.

Mercury is an environmental contaminant, which is highly toxic even at extremely low concentrations. Long-term accumulation of mercury in human body will damage the central nervous system or digestive tract system. Here, a new fluorescent chemical sensor Dansyl-His-Pro-Gly-Asp-NH2 (D-P4) was synthesized for the determination of Hg2+. The D-P4 sensor exhibits excellent selectivity and sensitivity to Hg2+ in aqueous solution with a 'turn-off' fluorescence response. Furthermore, D-P4-Hg system displays a good 'turn-on' fluorescence response to biothiols. The calculated binding constant for the 1:1 complex of D-P4 with Hg2 + is 1.07 × 105 M-1, which also confirms the high affinity of D-P4 for Hg2+. Results indicate that the detection limit of D-P4 for Hg2+ is 61.0 nM, and that of D-P4-Hg system for Cys is as low as 80.0 nM.