Dual-channel biosensor for simultaneous detection of S. typhimurium and L. monocytogenes using nanotags of gold nanoparticles loaded metal-organic frameworks.

Simultaneous detection of multiple foodborne pathogens is of great importance for ensuring food safety. Herein, we present a sensitive dual-channel electrochemical biosensor based on copper metal organic frameworks (CuMOF) and lead metal organic framework (PbMOF) for simultaneous detection of Salmonella typhimurium (S. typhimurium) and Listeria monocytogenes (L. monocytogenes). The MOF-based nanotags were prepared by functionalizing gold nanoparticles loaded CuMOF (Au@CuMOF) and PbMOF (Au@PbMOF) with signal DNA sequences 1 (sDNA1) and sDNA2, respectively. By selecting invA of S. typhimurium and inlA gene of L. monocytogenes as targe sequences, a sandwich-typed dual-channel biosensor was developed on glassy carbon electrodes (GCE) through hybridization reactions. The sensitive detection of S. typhimurium and L. monocytogenes was achieved by the direct differential pulse voltametric (DPV) signals of Cu2+ and Pb2+. Under optimal conditions, channel 1 of the biosensor showed linear range for invA gene of S. typhimurium in 1 × 10-14-1 × 10-8 M with low detection limit (LOD) of 3.42 × 10-16 M (S/N = 3), and channel 2 of the biosensor showed linear range for inlA gene of L. monocytogenes in 1 × 10-13-1 × 10-8 M with LOD of 6.11 × 10-15 M (S/N = 3). The dual-channel biosensor showed good selectivity which were used to detect S. typhimurium with linear range of 5-1.0 × 104 CFU mL-1 (LOD of 2.33 CFU mL-1), and L. monocytogenes with linear range of 10 - 1.0 × 104 CFU mL-1 (LOD of 6.61 CFU mL-1).

Real-Time Monitoring of a Single Molecule in Sub-nanometer Space by Dynamic Surface-Enhanced Raman Spectroscopy.

In biology and chemistry, the ultimate goal is to monitor single molecules without labels. However, long-term monitoring of label-free molecules remains a challenge. Here, on the basis of the photothermal effect of gold nanorods (GNRs), we developed a platform for monitoring of a single molecule employing surface-enhanced Raman spectroscopy (SERS). Laser re-irradiation forms 1.0 nm gaps between GNRs, allowing us to observe single crystal violet (CV) molecules blinking for up to 4 min with dynamic surface-enhanced Raman spectroscopy (D-SERS). Bianalyte experiments confirm single-molecule features at CV concentrations of 10-14 M. Combining density functional theory (DFT) calculations with a free CV molecule observed in millisecond D-SERS, we propose that CV molecules can be confined to sub-nanometer space and the orientation of an individual CV moving in the range of 50-90° can be dynamically captured by D-SERS. This will provide a novel idea for effective exploration of the temporal and spatial dynamic processes of different reactions.

Colorimetric biosensing of nopaline synthase terminator using Fe3O4@Au and hemin-functionalized reduced graphene oxide.

In this paper, we present a simple and label-free colorimetric biosensor for detection of the nopaline synthase (NOS) terminator in genetically modified (GM) plants. The "signal on" colorimetric biosensor was developed using a nanocomposite consisted of gold nanoparticles doped magnetic Fe3O4 nanoparticles (Fe3O4@Au NP), capture probe DNA (cDNA), and hemin-functionalized reduced graphene oxide nanosheets (H-GN). The nanocomposite was successfully prepared by means of Au-S bonds and the strong π interactions between cDNA and H-GN. The sensing approach is based on the excellent peroxidase-mimicking activity of H-GN and its different electrostatic interactions with single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA). In presence of the target NOS, the cDNA in the nanocomposite will hybridize with its complementary sequence, and form dsDNA structure. Due to the weak π interactions between dsDNA and H-GN, a portion of H-GN will be released from the surface of Fe3O4@Au NPs and transferred into solution. After magnetic separation was performed, the supernatant was incubated with 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of H2O2. The released H-GN can catalyze the oxidation reaction of TMB and turn the colorless solution blue. This "signal-on" colorimetric biosensor shows a broad linear range of 0.5-100 nM for the target NOS, with a 0.19 nM detection limit. The application of the biosensor for determination of NOS segments in samples of GM and non-GM tomatoes shows that it can discriminate between GM and non-GM plants. The reliability of the method for samples of NOS-spiked GM tomato suggests satisfactory recoveries in the range of 93.6%-94.2%.

Calcium ion assisted fluorescence determination of microRNA-167 using carbon dots-labeled probe DNA and polydopamine-coated Fe3O4 nanoparticles.

A selective and sensitive fluorescence biosensor is described for determination of microRNA-167 using fluorescent resonant energy transfer (FRET) strategy. The FRET system comprises carbon dots (CDs, donor) labeled with probe DNA (pDNA) and polydopamine (PDA)-coated Fe3O4 nanoparticles (Fe3O4@PDA NPs, acceptor). The CDs-pDNA can be absorbed onto the surface of Fe3O4@PDA NPs because of the strong π interaction between pDNA and PDA. With the enhanced adsorption ability of Fe3O4@PDA NPs by Ca2+, the fluorescence intensity of CDs at 445 nm (excitation at 360 nm) is quenched. In presence of microRNA-167, the hybridized complex of CDs-pDNA-microRNA-167 will be released from the surface of Fe3O4@PDA NPs due to the weak π interaction of the complex and PDA. This results in the fluorescence recovery of CDs. By application of twice-magnetic separation, the biosensor shows a wide linear range of 0.5-100 nM to microRNA-167 with a 76 pM detection limit. The method was applied to the determination of microRNA-167 in samples of total microRNA extractions from A. thaliana seedlings, and the recoveries ranged from 96.4 to 98.3%.

Electrochemical detection of Salmonella using an invA genosensor on polypyrrole-reduced graphene oxide modified glassy carbon electrode and AuNPs-horseradish peroxidase-streptavidin as nanotag.

A rapid and sensitive electrochemical biosensor was constructed to detect Salmonella using invA gene biosensor. The biosensing was based on polyrrole-reduced graphene oxide (PPy-rGO) nanocomposite modified glassy carbon electrode (GCE) and signal amplification with horseradish peroxidase-streptavidin biofunctionalized gold nanoparticles (AuNPs-HRP-SA). PPy-rGO was prepared at 60 °C by chemical reduction of PPy-functionalized graphene oxide (PPy-GO) that was synthesized by in situ polymerization at room temperature. The detection signal was amplified via enzymatic reduction of H2O2 in the presence of hydroquinone (HQ) using AuNPs-HRP-SA as nanotag. Under optimal conditions, the differential pulse voltametric (DPV) signal from the biosensor was linearly related to the logarithm of target invA gene concentrations from 1.0 × 10-16 to 1.0 × 10-10 M, and the limit of detection (LOD) was 4.7 × 10-17 M. The biosensor can also detect Salmonella in the range of 9.6 to 9.6 × 104 CFU mL-1, with LOD of 8.07 CFU mL-1. The biosensor showed good regeneration ability, acceptable selectivity, repeatability and stability, which bode well as an alternative method for Salmonella screening.