Reagentless detection of staphylococcal enterotoxin B via electrochemical interrogation of conformational changes.

An electrochemical biosensor for staphylococcal enterotoxin B (SEB) detection has been designed on the basis of electrochemical interrogation of conformational changes. Ferrocene-labeled hairpin probe (Fc-HP) and SEB aptamer are introduced for the construction of the platform. Without SEB, the rigid construction of DNA duplex that included SEB aptamer and Fc-HP prevented Fc getting access to the electrode surface, keeping the "eT-off" state in the detection system. In the presence of SEB, the interaction between SEB and the aptamer could trigger the disruption of DNA duplex and the restoration of hairpin structure, accompanied by the increase of Fc oxidation current. The decreasing distance between the redox probe and electrode upon the nucleic acid reconfiguration substantially increased the efficiency of eT, which resulted in the enhanced Fc signal. The proposed strategy presented a wide linear detection range from 0.005 to 100 ng mL-1 with a detection limit down to 3 pg mL-1 (S/N = 3). To investigate the applicability and reliability of the method in real food samples such as milk samples, we compared the results between this method and the commercial ELISA kit. The relative percentage error between the two assays ranged from -6.42% to 6.31%, indicating that there was no obvious difference between the results.

Chronocoulometric aptamer based assay for staphylococcal enterotoxin B by target-triggered assembly of nanostructured dendritic nucleic acids on a gold electrode.

A rapid and ultrasensitive method is described for the detection of staphylococcal enterotoxin B (SEB). It is based on the formation of a dendritic DNA superstructure by integrating (a) target-induced triggering of DNA release with (b) signal amplification by a hybridization chain reaction. Partially complementary pairing of aptamer and trigger DNA forms a duplex structure. The capture DNA is then placed on the surface of a gold electrode through gold-thiol chemistry. In the presence of SEB, the aptamer-target conjugate is compelled to form. This causes the release of trigger DNA owing to a strong competition with SEB. The trigger DNA is subsequently hybridized with the partial complementary sequences of the capture DNA to trigger HCR with three auxiliary DNA sequances (referred to as H1, H2, H3). Finally, the dendritic DNA superstructure is bound to hexaammineruthenium(III) cation by electrostatic adsorption and assembled onto the modified gold electrode. This produces an amplified electrochemical signal that is measured by chronocoulometry. Under optimal conditions, the charge difference increases linearly with the logarithm of the SEB concentrations in the range from 5 pg·mL-1 to 100 ng·mL-1 with a detection limit as low as 3 pg·mL-1 (at S/N = 3). Graphical abstract An electrochemical switching strategy is presented for the sensitive detection of Staphylococcus enterotoxin B based on target-triggered assembly of dendritic nucleic acid nanostructures.

Impedimetric determination of Staphylococcal enterotoxin B using electrochemical switching with DNA triangular pyramid frustum nanostructure.

An electrochemical switching strategy is presented for the sensitive determination of Staphylococcus enterotoxin B (SEB). It is based on the use of DNA triangular pyramid frustum nanostructure (TPFDNA) consisting of (a) three thiolated probes, (b) one auxiliary probe, and (c) an aptamer against SEB. The TPFDNA was assembled on the gold electrode, with the SEB aptamer designed on top of the TPFDNA. The electron transfer to hexacyanoferrate acting as an electrochemical probe is strongly inhibited in the TPFDNA-modified electrode. This is assumed to be due to the formation of a 3D TPFDNA structure that limits access of hexacyanoferrate to the electrode. Therefore, the Faradaic impedance is large. However, in the presence of SEB, it will bind to the aptamer and dehybridize the hybrid formed between aptamer and its complementary sequence. As a result, the TPFDNA nanostructure changes to an equilateral triangle DNA nanostructure. This results in a more efficient electron transfer and a smaller Faradaic impedance. The method has a detection limit of 0.17 ng mL-1 of SEB (at an S/N of 3) and a dynamic range that covers the 0.2-1000 ng mL-1 concentration range. The applicability and reliability of the method was demonstrated by anayzing (spiked) milk samples, and the results were compared to those obtained with an ELISA kit. The relative standard deviations between the two methods range between -6.59 and 9.33%. Graphical abstract An electrochemical switching strategy is presented for the sensitive detection of Staphylococcus enterotoxin B based on 3D DNA structure conversion of nanostructure from triangular pyramid frustum to equilateral triangle.

Application of Spectral Crosstalk Correction for Improving Multiplexed MicroRNA Detection Using a Single Excitation Wavelength.

MicroRNAs (miRNAs) play crucial roles in the regulation of cellular activities and are next-generation biomarkers for early cancer detection. Simultaneous monitoring of multiplexed miRNA is very important for enhancing the accuracy of cancer diagnostics. Traditional fluorescence methods for multicomponent analysis were usually operated under multiple excitation wavelengths, because spectral crosstalk is very detrimental to detecting accuracy for multicomponent analysis. Herein, we present a fluorescence strategy for multi-miRNAs detection in plasma under a single excitation wavelength. Nucleic acid stain TOTO-1 and three labeled fluorescence dyes Cy3, Cy3.5, and Cy5 emit no fluorescence in their free state. Target miRNA hybridized the auxiliary and probe oligonucleotides into duplex nucleic acid. Intercalation interaction localized TOTO-1 and labeled dyes into the duplex nucleic acid. As a result, TOTO-1 emitted strong fluorescence and efficient Förster resonance energy transfer (FRET) happened. MicroRNAs miRNA-155, miRNA-182, and miRNA-197, which are significant for the early diagnosis of lung cancer, were simultaneously detected as models. Deviations from spectral crosstalk in the presence of other miRNAs were corrected by mathematical methods. Results demonstrated that, after spectra crosstalk corrections, every miRNA at high or low concentration in plasma was determined accurately in the presence of either high or low concentrations of the other two miRNAs. This new multiplexed assay for miRNAs is promising for clinical diagnosis, prognosis, and therapeutic monitoring of early-stage lung cancer.

Visual, Label-Free Telomerase Activity Monitor via Enzymatic Etching of Gold Nanorods.

Early diagnosis and life-long surveillance are clinically important to improve the long-term survival of cancer patients. Telomerase activity is a valuable biomarker for cancer diagnosis, but its measurement often used complex label procedures. Herein, we designed a novel, simple, visual and label-free method for telomerase detection by using enzymatic etching of gold nanorods (GNRs). First, repeating (TTAGGG)x sequences were extented on telomerase substrate (TS) primer. It formed G-quadruplex under the help of Hemin and K+. Second, the obtained horseradish peroxidase mimicking hemin/G-quadruplex catalyzed the H2O2-mediated etching of GNRs to the short GNRs, even to gold nanoparticles (GNPs), generating a series of distinct color changes due to their plasmon-related optical response. Thus, this enzymatic reaction can be easily coupled to telomerase activity, allowing for the detection of telomerase activity based on vivid colors. This can be differentiated sensitively by naked eyes because human eyes are more sensitive to color variations rather than the optical density variations. As a result, telomerase activity can be quantitatively detected ranging from 200 to 15000 HeLa cells mL-1. The detection limit was 90 HeLa cells mL-1 (S/N = 3). Importantly, the application of this method in bladder cancer samples was in agreement with the clinical results. Thus, this method was considerably suitable for point-of-care diagnostics in resource-constrained regions because of the easy readout of results without the use of sophisticated apparatus.

In Situ Detection and Imaging of Telomerase Activity in Cancer Cell Lines via Disassembly of Plasmonic Core-Satellites Nanostructured Probe.

The label-free localized surface plasmon resonance (LSPR) detection technique has been identified as a powerful means for in situ investigation of biological processes and localized chemical reactions at single particle level with high spatial and temporal resolution. Herein, a core-satellites assembled nanostructure of Au50@Au13 was designed for in situ detection and intracellular imaging of telomerase activity by combining plasmonic resonance Rayleigh scattering spectroscopy with dark-field microscope (DFM). The Au50@Au13 was fabricated by using 50 nm gold nanoparticles (Au50) as core and 13 nm gold nanoparticles (Au13) as satellites, both of them were functionalized with single chain DNA and gathered proximity through the highly specific DNA hybridization with a nicked hairpin DNA (O1) containing a telomerase substrate (TS) primer as linker. In the presence of telomerase, the telomeric repeated sequence of (TTAGGG)n extended at the 3'-end of O1 would hybridized with its complementary sequences at 5'-ends. This led the telomerase extension product of O1 be folded to form a rigid hairpin structure. As a result, the Au50@Au13 was disassembled with the releasing of O1 and Au13-S from Au50-L, which dramatically decreased the plasmon coupling effect. The remarkable LSPR spectral shift was observed accompanied by a detectable color change from orange to green with the increase of telomerase activity at single particle level with a detection limit of 1.3 × 10-13 IU. The ability of Au50@Au13 for in situ imaging intracellular telomerase activity, distinguishing cancer cells from normal cells, in situ monitoring the variation of cellular telomerase activity after treated with drugs were also demonstrated.

Label-Free Detection of Telomerase Activity in Urine Using Telomerase-Responsive Porous Anodic Alumina Nanochannels.

Telomerase is closely related to cancers, which makes it one of the most widely known tumor marker. Recently, many methods have been reported for telomerase activity measurement in which complex label procedures were commonly used. In this paper, a label-free method for detection of telomerase activity in urine based on steric hindrance changes induced by confinement geometry in the porous anodic alumina (PAA) nanochannels was proposed. Telomerase substrate (TS) primer was first assembled on the inside wall of PAA nanochannels by Schiff reaction under mild conditions. Then, under the action of telomerase, TS primer was amplified and extended to repeating G-rich sequences (TTAGGG)x, which formed multiplex G-quadruplex in the presence of potassium ions (K(+)). This configurational change led to the increment of steric hindrance in the nanochannels, resulting in the decrement of anodic current of potassium ferricyanide (K3[Fe(CN)6]). Compared with previously reported methods based on PAA nanochannels (usually one G-quadruplex formed), multiplex repeating G-quadruplex formed on one TS primer in this work. As a result, large current drop (∼3.6 µA, 36%) was obtained, which gave facility to improve the detection sensitivity. The decreased ratio of anodic current has a linear correlation with the logarithm of HeLa cell number in the range of 10-5000 cells, with the detection limit of seven cells. The method is simple, reliable, and has been successfully applied in the detection of telomerase in urine with good accuracy, selectivity and reproducibility. In addition, the method is nondestructive test compared to blood analysis and pathology tests, which is significant for cancer discovery, development, and prognosis.

Chiroplasmonic Assemblies of Gold Nanoparticles for Ultrasensitive Detection of 8-Hydroxy-2'-deoxyguanosine in Human Serum Sample.

Gold nanoparticles (AuNPs) have been extensively explored to be used in analytical methods such as electrochemical, colorimetric methods, and so on. However, only a few methods have been reported by using chirality of AuNPs although their chiral assembly has been studied extensively and circular dichroism (CD) spectroscopy is also a simple and sensitive analytical method. In this paper, sensitive CD spectroscopy method has been explored for detection of 8-hydroxy-2'-deoxyguanosine (8-OHdG), a well-known biomarker for oxidative DNA damage, based on DNA-induced chiroplasmonic assemblies of AuNPs. First, 8-OHdG aptamer hybridized with its complementary sequence that modified with AuNPs based on precision matched bases. DNA-modified AuNPs were assembled into AuNPs dimers by 8-OHdG aptamer, which displayed strong chiroptical activity. Subsequently, in the presence of 8-OHdG, the high specific recognition and affinity constants of aptamer and 8-OHdG destroyed the hybrid of aptamer and its complementary sequence; as a result, AuNPs dimers were destroyed and showed low CD signal. The CD intensity was in log-linear correlation with the concentration of 8-OHdG ranging from 0.05 to 2 nM, with a correlation coefficient of 0.9951 and a detection limit of 33 pM (S/N = 3). The method has been successfully applied in a complex matrix such as human serum samples. The recoveries were from 92.5% to 107% and the relative standard derivations were in the range of 4.89% ∼ 7.27%, indicating that the method had good accuracy and high precision. Therefore, these results indicated that the proposed CD method was simple and reliable, which held great potential for clinical examinations.

Strategy for realizing magnetic field enhancement based on diffraction coupling of magnetic plasmon resonances in embedded metamaterials.

We have demonstrated a straightforward strategy to realize magnetic field enhancement through diffraction coupling of magnetic plasmon (MP) resonances by embedding the metamaterials consisting of a planar rectangular array of U-shaped metallic split-ring resonators (SRRs) into the substrate. Our method provides a more homogeneous dielectric background allowing stronger diffraction coupling of MP resonances among SRRs leading to strong suppression of the radiative damping. We observe that compared to the on-substrate metamaterials, the embedded ones lead to a narrow-band hybridized MP mode, which results from the interference between MP resonances in individual SRRs and an in-plane propagating collective surface mode arising from light diffraction. Associated with the excitation of this hybridized MP mode, a twenty-seven times enhancement of magnetic fields within the inner area of the SRRs is achieved as compared with the pure MP resonance. Moreover, we also found that besides the above requirement of homogeneous dielectric background, only a collective surface mode with its magnetic field of the same direction as the induced magnetic moment in the SRRs could mediate the excitation of such a hybridized MP mode.

Colorimetric detection of influenza A virus using antibody-functionalized gold nanoparticles.

Early and accurate diagnosis is considered the key issue to prevent the further spread of viruses and facilitate influenza therapy. Herein, we report a colorimetric immunosensor for influenza A virus (IAV) based on gold nanoparticles (AuNPs) modified with monoclonal anti-hemagglutinin antibody (mAb). The immunosensor allows for a fast, simple, and selective detection of IAV. In this assay, influenza-specific antibodies are conjugated to AuNPs to create mAb-AuNP probes. Since IAV has multiple recognition sites for probes on the surface, the mAb-AuNP probes can be specifically arranged on the virus surface due to their very specific antigen recognition. In this case, this aggregation of the mAb-AuNP probes produces a red shift in the absorption spectrum due to plasmon coupling between adjacent AuNPs, and it can be detected with the naked eye as a color change from red to purple and quantified with the absorption spectral measurements. The aggregate formation is also confirmed with transmission electron microscopy (TEM) imaging and dynamic light scattering (DLS). Under the optimal conditions, the present immunoassay can sensitively measure H3N2 IAV (A/Brisbane/10/2007) with a detection limit of 7.8 hemagglutination units (HAU). This proposed immunosensor revealed high specificity, accuracy, and good stability. Notably, it is a single-step detection using AuNP probes and UV-vis spectrophotometer for readout, and no additional amplification, e.g., enzymatic, is needed to read the result. This assay depends on an ordered AuNP structure covering the virus surface and can be applied to any virus pathogen by incorporating the appropriate pathogen-specific antibody.

Label-free fluorescence detection of DNA methylation and methyltransferase activity based on restriction endonuclease HpaII and exonuclease III.

Strategies to detect the methylation of site specific DNA and assay of M.SssI methyltransferase (M.SssI MTase) activity are important in determining human cancers due to aberrant methylation linked to cancer initiation and progression. Herein, we report a label-free fluorescence detection method for DNA methylation and MTase activity based on restriction endonuclease HpaII and exonuclease III (Exo III). A label-free probe DNA was designed, which hybridized with target DNA (one 32-mer DNA from the exon 8 promoter region of the Homo sapiens p53 gene) to form double stranded DNA (dsDNA). Upon the cleavage action of HpaII and degradation reaction of Exo III, dsDNA changed to single stranded DNA (ssDNA) and the fluorescence intensity of thiazole orange (TO) is weak. After the resulting dsDNA was methylated by M.SssI MTase, the action of HpaII and Exo III was prevented, then TO intercalates into the dsDNA and emits strong fluorescence. This method can determine DNA methylation at the site of CpG and distinguish a one-base mismatched target sequence. The fluorescence intensity has a linear relationship with M.SssI MTase activities in the range of 1-10 U mL(-1) with a detection limit of 0.16 U mL(-1) in terms of 3 times deviation of the blank sample. The methylation of DNA by a hydroxyl radical triggered by DMSO and CH3CHO was also measured. These results show that the proposed method can specifically and selectively detect DNA methylation and M.SssI MTase activity. Human serum has no obvious effects on the assay performance, indicating that the method has great potential for further application in complex samples.

Rapid screening of electrochemically active bacteria based on a biocathode-functional bipolar electrode-electrochemiluminescence platform.

A functional bipolar electrode-electrochemiluminescence (BPE-ECL) platform based on biocathode reducing oxygen was constructed for detecting electrochemically active bacteria (EAB) in this paper. Firstly, thiolated trimethylated chitosan quaternary ammonium salt (TMC-SH) layer was assembled on the gold-plated cathode of BPE. TMC-SH contains quaternary ammonium salt branch chain, which can inhibit the growth of microorganisms on the surface or in the surrounding environment while absorbing bacteria. Then, the peristaltic pump was used to flow all of the samples through the cathode, and the EAB was electrostatically adsorbed on the electrode surface. Finally, applying a constant potential to the BPE, bacteria can catalyze electrochemical reduction of O2, and decrease the overpotential of O2 reduction at the cathode, which in turn generates an ECL reporting intensity change at the anode. In this way, live and dead bacteria can be distinguished, and the influence of complex food substrates on detection can be greatly reduced.

Biocathodes reducing oxygen in BPE-ECL system for rapid screening of E. coli O157:H7.

After discovery of electron transfer from bacteria, most bacteria known to be electrochemically active are utilized as a self-regenerable catalyst at the anode of microbial fuel cells (MFCs). However, the reverse phenomenon, cathodic catalysts is not so widely researched. This present study demonstrated that E. coli O157:H7 was electrochemically active, and it was able to catalyze oxygen reduction at the cathode of bipolar electrode (BPE). Applying a constant potential to the BPE, E. coli O157:H7 can catalyze electrochemical reduction of O2, decrease the overpotential of O2 reduction at the cathode, which in turn generates an electrochemiluminescence (ECL) reporting intensity change at the anode. Significantly, a majority of food matrix does not exhibit catalytic activity for electrochemical reduction of O2. Meanwhile, due to the physically separation of two poles of closed BPE, complex food matrix at the cathode does not interfere with the ECL reaction at the anode. Therefore, the effect of food matrix is negligible when measuring E. coli O157:H7 levels in food. A low detection limit of 10 CFU mL-1 E. coli O157:H7 could be identified within 1 h. Thus, biocathodes reducing oxygen in BPE-ECL system has shown excellent characteristics in the field of rapid detection of electroactive bacteria in food.

Double bipolar electrode electrochemiluminescence color switch for food-borne pathogens detection.

Color-switch electrochemiluminescence (ECL) sensing platform based on a dual-bipolar electrode (D-BPE) is reported in this work. The D-BPE was composed of a cathode filled with buffer and two anodes filled with [Ru(bpy)3]2+-TPrA and luminol-H2O2 solutions, respectively. Both anodes were modified with capture DNA and served as ECL reporting platforms. After introducing ferrocene-labeled aptamer (Fc-aptamer) on both anodes, the ECL emission signal of the [Ru(bpy)3]2+ was difficult to be observed (anode 1), while luminol emitted a strong and visible ECL signal (anode 2). Ferrocene (Fc) did not only prevent the oxidation of [Ru(bpy)3]2+ due to its lower oxidation potential, its oxidation product Fc+ also quenched the [Ru(bpy)3]2+ ECL through efficient energy transfer. For luminol, Fc+ catalyzes the accelerated formation of the excited-state of the luminol anion radical, which leads to the enhancement of the luminol ECL. In the presence of food-borne pathogens, the aptamer was assembled with them, leading to the leaving of Fc from the surface of the D-BPE anodes. The ECL intensity of [Ru(bpy)3]2+ was enlarged, meanwhile, the blue emission signal of luminol became weakened. By self-calibrating the ratio of the two signals, 1-106 CFU mL-1 food-borne pathogenic bacteria can be sensitively detected with a detection limit of 1 CFU mL-1. Ingeniously, the color-switch biosensor can be used to detect S. aureus, E. coli and S. typhimurium by assembling the corresponding aptamers onto the D-BPE anodes.

Temporal sensing platform based on anodic dissolution of Ag and cathodic biocatalysis of oxygen reduction for Staphylococcus aureus detection.

An Ag@C hybrid bipolar electrode (BPE) sensing platform has been established for the temporal detection of Staphylococcus aureus (S. aureus) in food. Combining the advantages of anodic dissolution of Ag and cathodic biocatalysis of oxygen (O2) reduction, this strategy showed an ultralow detection limit down to 10 CFU mL-1. As the formation of Ag@C completely quenched the electrochemiluminescence (ECL) emission of luminol, the ECL emission recovery reflected the extent of anodic dissolution. Meanwhile, S. aureus catalyzed the electrochemical reduction of O2 at the cathode, reducing the overpotential for cathodic O2 reduction and thus increasing the rate of anodic electron loss, facilitating Ag dissolution and restoring the ECL emission of luminol. When a constant potential was applied, through monitoring the ECL recovery time before and after the incubation of S. aureus on the cathode, S. aureus could be quantified due to the slight difference of the conductivity.

One-step grain pretreatment for ochratoxin A detection based on bipolar electrode-electrochemiluminescence biosensor.

False positives are common and frequently occurring in detection of ochratoxin A (OTA) due to the complexity of the food matrix. In this paper, a novel bipolar electrode-electrochemiluminescence (BPE-ECL) sensing platform for sensitive OTA detection with one-step grain pretreatment was proposed. The biosensor uses cathode of closed BPE as a functional sensing interface and anode as a signal collection interface. On the functional sensing interface, the horseradish peroxidase (HRP) catalyze the polymerization of aniline to form polyaniline (PANI) on nucleic acid backbone which is supplied by DNA tetrahedron-structured aptamer (DTA) and hybrid chain reaction (HCR). In the presence of OTA, PANI is formed and can cause the change of ECL and luminescence voltage of the anode of BPE. On the signal collection interface, the Ru(bpy)32+/TPA system is used as ECL light output. In this way, the analyte does not need to participate the ECL reaction of the anode, which avoids direct contact of photoactive molecules with complex reaction systems and greatly reduce the influence of complex food matrix on signal acquisition. The accuracy of the BPE-ECL biosensor (one-step grain pretreatment) was similar with high performance liquid chromatography (HPLC) analysis (traditional national standard pretreatment method: GB5009.96-2016). Meanwhile, the BPE-ECL biosensor had higher sensitivity (LOD: 3 pg mL-1). Therefore, closed BPE could simplify sample pretreatment and improve detection capability.

Simultaneous fluorescent detection of multiplexed miRNA of liver cancer based on DNA tetrahedron nanotags.

The level of miRNA-21, miRNA-122, and miRNA-223 are always elevated when liver cancer is present at an early stage. In this paper, a novel assay to simultaneous detect miRNA-21, miRNA-122, and miRNA-223 was proposed based on DNA tetrahedron nanotags and fluorescence resonance energy transfer (FRET), which used a single laser stimulate wavelengh from one nucleic acid stain TOTO-1 to three diverse organic dyes (Cy3, Cy3.5, Cy5). In brief, a DNA tetrahedral nanostructure (DTN) was designed with three adaptor oligos on its vertices. TOTO-1, as a fluorescent donor, can imbed into native nucleic acid backbone of DTN. Three organic dye-functionalized strands (FRET oligos) are fluorescent receptors. In the presence of target miRNAs, they can be hybridized with FRET oligos and adaptor oligos on the vertices of DTN and the stable DNA tetrahedron nanotags are formed. As a result, the confinement of TOTO-1 is in close proximity to three fluorescence dyes, the FRET between TOTO-1 and three fluorescence dyes is generate efficiently in DNA tetrahedron nanotags. Point-of-care clinical applicability is demonstrated by sensitive multiplexed quantification of three miRNAs in 10% human serum samples.

Development of a real-time PCR assay for the identification and quantification of bovine ingredient in processed meat products.

In order to find fraudulent species substitution in meat products, a highly sensitive and rapid assay for meat species identification and quantification is urgently needed. In this study, species-specific primers and probes were designed from the mitochondrial cytb (cytochrome b) fragment for identification and quantification of bovine ingredient in commercial meat products. Bovine samples and non-bovine ones were used to identify the specificity, sensitivity, and applicability of established assay. Results showed that the primers and probes were highly specific for bovine ingredient in meat products. The absolute detection limit of the real-time PCR method was 0.025 ng DNA, and the relative detection limit was 0.002% (w/w) of positive samples. The quantitative real-time PCR assay was validated on simulated meat samples and high in the precision and accuracy. In order to demonstrate the applicability and reliability of the proposed assay in practical products, the 22 commercial meat products including salted, jerkies, and meatball were used. The results indicated the established method has a good stability in detection of bovine ingredient in real food. The established method in this study showed specificity and sensitivity in identification and quantification of beef meat in processed meat products.

Fluorescent difference between two rhodamine-PAHs polystyrene solid-phase sensors for Hg(II) detection based on crystal structure and density functional theory calculation.

Two novel rhodamine-polystyrene solid-phase fluorescence sensors PS-RB-2 and PS-R6G-2 with pyrene or naphthalene as fluorophore were synthesized for Hg(II) detection. Their structures were characterized by Fourier transform infrared (FTIR) spectra and scanning electron micrographs (SEM). Sensor PS-RB-2 displayed higher selectivity and sensitivity to Hg(II), with a lower detection limit of 0.065 µM. A detection mechanism involving the Hg(II) chelation-induced spirocycle open of rhodamine was proposed and discussed from theoretic level based on crystal structures and density functional theory (DFT) calculations. Sensor PS-RB-2 with recyclable and environment-friendly performance was successfully employed to fluorescent detection of Hg(II) in real water and fish samples, indicating its good potential in practical application. Its solid phase extraction columns were developed for rapid detection of Hg(II) by observing the color change with the naked eyes.

Sensitive electrochemical detection of aflatoxin B1 using DNA tetrahedron-nanostructure as substrate of antibody ordered assembly and template of aniline polymerization.

A novel strategy for AFB1 detection in grains was proposed based on DNA tetrahedron-structured probe (DTP) and horseradish peroxidase (HRP) triggered polyaniline (PANI) deposition. Briefly, the DNA tetrahedron nanostructures were assembled on the gold electrode, with carboxylic group designed on top vertex of them. The carboxylic group was conjugated with the AFB1 monoclonal antibody (mAb) to form DTP. The test sample and a known fixed concentration of HRP-labeled AFB1 were mixed and they compete for binding to DTP. The HRP assembled on the gold electrode catalyzed the polymerization of aniline on DTP. AFB1 in grains could be determined by using PANI as electrochemical signal molecules. Interestingly, DNA tetrahedron-structure, which has mechanical rigidity and structural stability, can improve antigen-antibody specific recognition and binding efficiency through the use of mAb ordered assembly. Meanwhile, nucleic acid backbone with a large amount of negative charge is good template for aniline polymerization under mild conditions.