Weiwei Ye

A fluorescence resonance energy transfer (FRET) biosensor based on graphene quantum dots (GQDs) and gold nanoparticles (AuNPs) for the detection of mecA gene sequence of Staphylococcus aureus

In this work, a novel fluorescence resonance energy transfer (FRET) biosensor based on graphene quantum dots (GQDs) and gold nanoparticles (AuNPs) pairs was developed for Staphylococcus aureus specific gene sequence detection. This FRET biosensor platform was realized by immobilization of capture probes on GQDs and conjugation of reporter probes on AuNPs. Target oligos then co-hybridized with capture probes and reporter probes to form a sandwich structure which brought GQDs and AuNPs to close proximity to trigger FRET effect. The fluorescence signals before and after addition of targets were measured and the fluorescence quenching efficiency could reach around 87% with 100 nM target oligo. The limit of detection (LOD) of this FRET biosensor was around 1 nM for S.aureus gene detection. Experiments with both single-base mismatched oligos and double-base mismatched oligos demonstrated the good sequence selectivity of this FRET biosensor.

Ultrasensitive Detection of Ebola Virus Oligonucleotide Based on Upconversion Nanoprobe/Nanoporous Membrane System

Ebola outbreaks are currently of great concern, and therefore, development of effective diagnosis methods is urgently needed. The key for lethal virus detection is high sensitivity, since early-stage detection of virus may increase the probability of survival. Here, we propose a luminescence scheme of assay consisting of BaGdF5:Yb/Er upconversion nanoparticles (UCNPs) conjugated with oligonucleotide probe and gold nanoparticles (AuNPs) linked with target Ebola virus oligonucleotide. As a proof of concept, a homogeneous assay was fabricated and tested, yielding a detection limit at picomolar level. The luminescence resonance energy transfer is ascribed to the spectral overlapping of upconversion luminescence and the absorption characteristics of AuNPs. Moreover, we anchored the UCNPs and AuNPs on a nanoporous alumina (NAAO) membrane to form a heterogeneous assay. Importantly, the detection limit was greatly improved, exhibiting a remarkable value at the femtomolar level. The enhancement is attributed to the increased light-matter interaction throughout the nanopore walls of the NAAO membrane. The specificity test suggested that the nanoprobes were specific to Ebola virus oligonucleotides. The strategy combining UCNPs, AuNPs, and NAAO membrane provides new insight into low-cost, rapid, and ultrasensitive detection of different diseases. Furthermore, we explored the feasibility of clinical application by using inactivated Ebola virus samples. The detection results showed great potential of our heterogeneous design for practical application.

Extracellular potentials recording in intact olfactory epithelium by microelectrode array for a bioelectronic nose

Human beings and animals have sensitive olfactory systems that can sense and identify a variety of odors. The purpose of this study is to combine biological cells with micro-chips to establish a novel bioelectronic nose system for odor detection by electrophysiological sensing measurements of olfactory tissue. In our experiments, 36-channel microelectrode arrays (MEAs) with the diameter of 30 microm were fabricated on the glass substrate, and olfactory epithelium was stripped from rats and fixed on the surface of MEA. Electrophysiological activities of olfactory receptor neurons in intact epithelium were measured through the multi-channel recording system. The extracellular potentials of cell networks could be effectively analyzed by correlation analysis between different channels. After being stimulated by odorants, such as acetic acid and butanedione, the olfactory cells generate different firing modes. These firing characteristics can be derived by time-domain and frequency-domain analysis, and they were different from spontaneous potentials. The investigation of olfactory epithelium can provide more information of olfactory system for artificial olfaction biomimetic design.

A graphene oxide based fluorescence resonance energy transfer (FRET) biosensor for ultrasensitive detection of botulinum neurotoxin A (BoNT/A) enzymatic activity

Botulinum neurotoxins (BoNTs) are among the most potent toxic bacterial proteins for humans, which make them potential agents for bioterrorism. Therefore, an ultrasensitive detection of BoNTs and their active states is in great need as field-deployable systems for anti-terrorism applications. We report the construction of a novel graphene oxide (GO)-peptide based fluorescence resonance energy transfer (FRET) biosensor for ultrasensitive detection of the BoNT serotype A light chain (BoNT-LcA) protease activity. A green fluorescence protein (GFP) modified SNAP-25 peptide substrate (SNAP-25-GFP) was optimally designed and synthesized with the centralized recognition/cleavage sites. This FRET platform was constructed by covalent immobilization of peptide substrate on GO with BSA passivation which have advantages of low non-specific adsorption and high stability in protein abundant solution. BoNT-LcA can specifically cleave SNAP-25-GFP substrate covalently immobilized on GO to release the fragment with GFP. Based on fluorescence signal recovery measurement, the target BoNT-LcA was detected sensitively and selectively with the linear detection range from 1fg/mL to 1pg/mL. The limit of detection (LOD) for BoNT-LcA is around 1fg/mL.

Olfactory receptor cells respond to odors in a tissue and semiconductor hybrid neuron chip

Olfactory systems of human beings and animals have the abilities to sense and distinguish varieties of odors. In this study, a bioelectronic nose was constructed by fixing biological tissues onto the surface of light-addressable potentiometric sensor (LAPS) to mimic human olfaction and realize odor differentiation. The odorant induced potentials on tissue-semiconductor interface was analyzed by sensory transduction theory and sheet conductor model. The extracellular potentials of the receptor cells in the olfactory epithelium were detected by LAPS. Being stimulated by different odorants, such as acetic acid and butanedione, olfactory epithelium activities were analyzed on basis of local field potentials and presented different firing modes. The signals fired in different odorants could be distinguished into different clusters by principal component analysis (PCA). Therefore, with cellular populations well preserved, the epithelium tissue and LAPS hybrid system will be a promising neuron chip of olfactory biosensors for odor detecting.

Extracellular recording of spatiotemporal patterning in response to odors in the olfactory epithelium by microelectrode arrays.

In olfactory biosensors, microelectronic sensor chips combined with biological olfactory cells are a promising platform for odor detection. In our investigation, olfactory epithelium stripped from rat was fixed on the surface of microelectrode arrays (MEAs). Electrophysiological activities of olfactory receptor neurons in intact epithelium were measured in the form of extracellular potentials. Based on multi-channel recording performance of MEA and structural and functional integrality of native olfactory epithelium, the spatiotemporal analysis was carried out to study the extracellular activity pattern of neurons in the tissue. The variation of spatiotemporal patterns corresponding to different odors displayed the signals firing image characteristic intuitionally. It is an effective method in the form of patterns for monitoring the state of tissue both in time and space domain, promoting the platform for olfactory sensing mechanism research.

Nanoporous membrane based impedance sensors to detect the enzymatic activity of botulinum neurotoxin A

Botulinum neurotoxins are among the most potent toxic bacterial proteins for humans and there is a great need to develop simple, rapid and sensitive methods for toxin detection and protease activity quantification in field deployment. In this paper, a nanoporous membrane based impedance sensor was developed to monitor the activity of the BoNT serotype A light chain protease (LcA). Synaptosomal-associated protein 25 (SNAP-25) was first immobilized inside nanopore walls via silane linkers. BoNT LcA was then injected over the nanoporous membrane substrate sensor and specifically cleaved SNAP-25. The cleavage activity could be monitored by measuring impedance signals across nanoporous membranes which represented the nanopore blockage degree. This initial device could achieve a 500 pM LcA detection limit within 25 minutes.

A hydrophilic polymer based microfluidic system with planar patch clamp electrode array for electrophysiological measurement from cells.

This paper presents a microfluidic planar patch clamp system based on a hydrophilic polymer poly(ethylene glycol) diacrylate (PEGDA) for whole cell current recording. The whole chip is fabricated by UV-assisted molding method for both microfluidic channel structure and planar electrode partition. This hydrophilic patch clamp chip has demonstrated a relatively high gigaseal success rate of 44% without surface modification compared with PDMS based patch clamp devices. This chip also shows a capability of rapid intracellular and extracellular solution exchange with high stability of gigaseals. The capillary flow kinetic experiments demonstrate that the flow rates of PEGDA microfluidic channels are around two orders of magnitude greater than those for PDMS-glass channels with the same channel dimensions. This hydrophilic polymer based patch clamp chips have significant advantages over current PDMS elastomer based systems such as no need for surface modification, much higher success rate of cell gigaseals and rapid solution exchange with stable cell gigaseals. Our results indicate the potential of these devices to serve as useful tools for pharmaceutical screening and biosensing tasks.

A Nanoporous Alumina Membrane Based Electrochemical Biosensor for Histamine Determination with Biofunctionalized Magnetic Nanoparticles Concentration and Signal Amplification.

Histamine is an indicator of food quality and indispensable in the efficient functioning of various physiological systems. Rapid and sensitive determination of histamine is urgently needed in food analysis and clinical diagnostics. Traditional histamine detection methods require qualified personnel, need complex operation processes, and are time-consuming. In this study, a biofunctionalized nanoporous alumina membrane based electrochemical biosensor with magnetic nanoparticles (MNPs) concentration and signal amplification was developed for histamine determination. Nanoporous alumina membranes were modified by anti-histamine antibody and integrated into polydimethylsiloxane (PDMS) chambers. The specific antibody modified MNPs were used to concentrate histamine from samples and transferred to the antibody modified nanoporous membrane. The MNPs conjugated to histamine were captured in the nanopores via specific reaction between histamine and anti-histamine antibody, resulting in a blocking effect that was amplified by MNPs in the nanopores. The blockage signals could be measured by electrochemical impedance spectroscopy across the nanoporous alumina membrane. The sensing platform had great sensitivity and the limit of detection (LOD) reached as low as 3 nM. This biosensor could be successfully applied for histamine determination in saury that was stored in frozen conditions for different hours, presenting a potentially novel, sensitive, and specific sensing system for food quality assessment and safety support.

Rapid and Sensitive Detection of Bacteria Response to Antibiotics Using Nanoporous Membrane and Graphene Quantum Dot (GQDs)-Based Electrochemical Biosensors

The wide abuse of antibiotics has accelerated bacterial multiresistance, which means there is a need to develop tools for rapid detection and characterization of bacterial response to antibiotics in the management of infections. In the study, an electrochemical biosensor based on nanoporous alumina membrane and graphene quantum dots (GQDs) was developed for bacterial response to antibiotics detection. Anti-Salmonella antibody was conjugated with amino-modified GQDs by glutaraldehyde and immobilized on silanized nanoporous alumina membranes for Salmonella bacteria capture. The impedance signals across nanoporous membranes could monitor the capture of bacteria on nanoporous membranes as well as bacterial response to antibiotics. This nanoporous membrane and GQD-based electrochemical biosensor achieved rapid detection of bacterial response to antibiotics within 30 min, and the detection limit could reach the pM level. It was capable of investigating the response of bacteria exposed to antibiotics much more rapidly and conveniently than traditional tools. The capability of studying the dynamic effects of antibiotics on bacteria has potential applications in the field of monitoring disease therapy, detecting comprehensive food safety hazards and even life in hostile environment.