Lanhua Liu

A reusable aptamer-based evanescent wave all-fiber biosensor for highly sensitive detection of Ochratoxin A.

Although aptamer-based biosensors have attracted ever-increasing attentions and found potential applications in a wide range of areas, they usually adopted the assay protocol of immobilizing DNA probe (e.g., aptamer, aptamer-complementary oligonucleotides) on a solid sensing surface, making it critical and challengeable to keep the integration of nucleic acid surface during the regeneration and the restoration to its original DNA probe form after repeated uses. In order to address the issue, we report a novel aptamer-based biosensing strategy based on an evanescent wave all-fiber (EWA) platform. In a simple target capturing step using aptamer-functionalized magnetic microbeads, signal probes conjugated with streptavidin are released and further detected by a EWA biosensor via a facial dethiobiotin-streptavidin recognition. Apart from the inherent advantages of aptamer-based evanescent wave biosensors (e.g. target versatility, sensitivity, selectivity and portability), the proposed strategy exhibits a high stability and remarkable reusability over other aptasensors. Under the optimized conditions, the typical calibration curve obtained for Ochratoxin A has a detection limit of 3nM with a linear response ranging from 6nM to 500nM. The dethiobiotin-streptavidin sensing surface instead of the traditional nucleic acid one can be reused for over 300 times without losing sensitivity.

Portable optical aptasensor for rapid detection of mycotoxin with a reversible ligand-grafted biosensing surface

As food safety is gaining prominence as a global issue, the demand for developing rapid, simple, on-site, accurate and low-cost biosensor technologies will continue to grow. This study demonstrates an evanescent wave optical aptasensor with a reversible ligand-grafted biosensing surface for rapid, sensitive and highly selective detection of ochratoxin A (OTA) in food. In this system, the OTA molecules were covalently immobilized onto the surface of an optical fiber using glutaraldehyde and ethylenediamine as space linkers. An integrated evanescent wave all-fiber (EWA) biosensing platform was developed for investigating the binding kinetics between the tethered ligand and free OTA-aptamer, the performance of the aptamer-based bioassay and the reversibility of biosensing surface. The affinity constant (Ka) of aptamer with tethered OTA was measured to be 2.2 × 10(8)M(-1) based on the EWA biosensing platform. With a competitive detection mode, the quantification of OTA over concentration ranges from 0.73 μg L(-1) to 12.50 μg L(-1) with a detection limit of 0.39 μg L(-1). The performance of the aptasensor with other interfering mycotoxins and spiked real wheat samples shows high specificity and selectivity, good recovery, precision, and accuracy, indicating that it can be applied for on-site, inexpensive and easy-to-use monitoring of OTA in real samples. Moreover, since the organic ligands are grafted onto the fiber surface, this strategy may avoid the potential disadvantages caused by immobilizing the nucleic acid biomolecules, such as weak restoration to the original DNA conformation after repeated uses.

Highly sensitive and selective optofluidics-based immunosensor for rapid assessment of Bisphenol A leaching risk.

Bisphenol A (BPA), a xenoestrogenic endocrine-disrupting chemical, is used in many consumer products worldwide and is widely detected in the environment and in food. Combining the advantages of evanescent wave fiber optic sensor and microfluidic technology, an all-fiber optofluidics-based bioassay platform (AFOB) was developed for the rapid immunoassay and assessment of BPA. The captured molecular BPA-bovine serum albumin was covalently immobilized on the surface of the fiber optic sensor. A mixture of different concentrations of BPA and a certain concentration of fluorescence-labeled anti-BPA monoclonal antibodies after pre-reaction was introduced to the optofluidic cell. A higher concentration of BPA reduced the fluorescence-labeled antibodies bound to the sensor surface and thus reduced fluorescence signals. Under optimal conditions, the BPA quantified as 0.5-100 μg/L, with a detection limit of 0.06 μg/L. The high selectivity of the sensor was evaluated in terms of its response to several potentially interfering chemicals. The potential interference of an environmental sample matrix was assessed by spiked samples, and the recovery of BPA ranged from 90% to 120% with relative standard deviation values of <9.1%. The AFOB and high-performance liquid chromatography had a desired correlation (R(2)=0.9958). The sensing platform was successfully used to assess BPA leaching from polycarbonate bottles at 45 °C and 80 °C, indicating that more BPA was substantially leached at elevated temperature and extend time. Thus, the developed sensing strategy can be an alternative method to rapidly analyze and assess the migration mechanism and fate of BPA or other pollutants.

Direct electron transfer reaction of laccase on a glassy carbon electrode modified with 1-aminopyrene functionalized reduced graphene oxide

A biosensor was developed based on the covalent immobilization of laccase (Lac) on 1-aminopyrene (1-AP)-functionalized reduced graphene oxide (rGO)-modified glassy carbon electrode (GCE) by encapsulation with chitosan. X-ray photoelectron spectroscopy and cyclic voltammetry were used to optimize the mass ratio between 1-AP and rGO (rAP/rGO) to be 1/8. Direct electron transfer behavior of Lac immobilized on 1-AP-functionalized rGO (AP-rGO) was investigated by cyclic voltammogram. Results indicate that the AP-rGO is a friendly platform for the immobilization of enzyme, which facilitates the electron exchange between Lac and the modified electrode surface. On AP-rGO/GCE, Lac displayed rapid electron transfer with one participating proton. Furthermore, it was also confirmed that Lac immobilized on the modified electrode exhibited the electrocatalytic response to an established unspecific Lac substrate, 2,2′-azinobis (3-ethylbenzothiazoline-6-sulfonate)diammonium salt (ABTS). Without the aid of an electron mediator between Lac and modified electrode, the enzyme-based biosensor shows good reproducibility and high sensitivity to ABTS with a detection limit of 0.1 μM (S/N = 3). Within the concentration of 2–1000 μM, a linear relationship was achieved between the current intensity of the electro-catalytic oxidation of ABTS and the concentration of ABTS. The proposed biosensor achieved 95% of steady-state reduction currents within 5 s.

Utilization of unmodified gold nanoparticles for label-free detection of mercury (II): Insight into rational design of mercury-specific oligonucleotides

Colorimetric detection of mercury (II) with the use of DNA oligonucleotides and unmodified gold nanoparticles (AuNPs) as indicators has been extensively studied. This study provides in-depth insights into the rational design of mercury-specific oligonucleotides (MSO) in the biosensing system. The leftover bases of MSO, as a result of the formation of T-Hg2+-T base pairs, can adsorb on the AuNPs and hinder their aggregation at concentrations of salt. This phenomenon was directly verified by the changes in particle sizes characterized by dynamic light scattering for the first time. Based on these findings, we proposed a rational design for the MSO with approximately 20-fold improvement in detection sensitivity. The detection limit of the proposed assay decreased to 15nM with a linear working range from 50nM to 300nM for Hg2+. The cross-reactivity against eight other metal ions was negligible compared with the response to Hg2+. Considering the diverse applications of AuNPs with oligonucleotides, this study can serve as a good reference and provides important implications in sensing and DNA-directed nanoparticle assembly.

Facile screening of potential xenoestrogens by an estrogen receptor-based reusable optical biosensor

The apparent increase in hormone-induced cancers and disorders of the reproductive tract has led to a growing demand for new technologies capable of screening xenoestrogens. We reported an estrogen receptor (ER)-based reusable fiber biosensor for facile screening estrogenic compounds in environment. The bioassay is based on the competition of xenoestrogens with 17β-estradiol (E2) for binding to the recombinant receptor of human estrogen receptor α (hERα) protein, leaving E2 free to bind to fluorophore-labeled anti-E2 monoclonal antibody. Unbound anti-E2 antibody then binds to the immobilized E2-protein conjugate on the fiber surface, and is detected by fluorescence emission induced by evanescent field. As expected, the stronger estrogenic activity of xenoestrogen would result in the weaker fluorescent signal. Three estrogen-agonist compounds, diethylstilbestrol (DES), 4-n-nonylphenol (NP) and 4-n-octylphenol (OP), were chosen as a paradigm for validation of this assay. The rank order of estrogenic potency determined by this biosensor was DES>OP>NP, which were consistent with the published results in numerous studies. Moreover, the E2-protein conjugate modified optical fiber was robust enough for over 300 sensing cycles with the signal recoveries ranging from 90% to 100%. In conclusion, the biosensor is reusable, reliable, portable and amenable to on-line operation, providing a facile, efficient and economical alternative to screen potential xenoestrogens in environment.

Integrated optical waveguide-based fluorescent immunosensor for fast and sensitive detection of microcystin-LR in lakes: optimization and analysis

Nowadays, biosensor technologies which can detect various contaminants in water quickly and cost-effectively are in great demand. Herein, we report an integrated channel waveguide-based fluorescent immunosensor with the ability to detect a maximum of 32 contaminants rapidly and simultaneously. In particular, we use waveguide tapers to improve the efficiency of excitation and collection of fluorescent signals in the presence of fluorophore photobleaching in a solid surface bioassay. Under the optimized waveguide geometry, this is the first demonstration of using such a type of waveguide immunosensor for the detection of microcystin-LR (MC-LR) in lake water. The waveguide chip was activated by (3-Mercaptopropyl) trimethoxysilane/N-(4-maleimidobutyryloxy) succinimide (MTS/GMBS) for immobilization of BSA-MC-LR conjugate, which was confirmed to have uniform monolayer distribution by atomic force microscopy. All real lake samples, even those containing MC-LR in the sub-microgram per liter range (e.g. 0.5 μg/L), could be determined by the immunosensor with recovery rates between 84% and 108%, confirming its application potential in the measurement of MC-LR in real water samples.

Isoelectric Bovine Serum Albumin: Robust Blocking Agent for Enhanced Performance in Optical-Fiber Based DNA Sensing

Surface blocking is a well-known process for reducing unwanted nonspecific adsorption in sensor fabrication, especially important in the emerging field where DNA/RNA applied. Bovine serum albumin (BSA) is one of the most popular blocking agents with an isoelectric point at pH 4.6. Although it is widely recognized that the adsorption of a blocking agent is strongly affected by its net charge and the maximum adsorption is often observed under its isoelectric form, BSA has long been perfunctorily used for blocking merely in neutral solution, showing poor blocking performances in the optical-fiber evanescent wave (OFEW) based sensing toward DNA target. To meet this challenge, we first put forward the view that isoelectric BSA (iep-BSA) has the best blocking performance and use an OFEW sensor platform to demonstrate this concept. An optical-fiber was covalently modified with amino-DNA, and further coupled with the optical system to detect fluorophore labeled complementary DNA within the evanescent field. A dramatic improvement in the reusability of this DNA modified sensing surface was achieved with 120 stable detection cycles, which ensured accurate quantitative bioassay. As expected, the iep-BSA blocked OFEW system showed enhanced sensing performance toward target DNA with a detection limit of 125 pM. To the best of our knowledge, this is the highest number of regeneration cycles ever reported for a DNA immobilized optical-fiber surface. This study can also serve as a good reference and provide important implications for developing similar DNA-directed surface biosensors.

Highly sensitive detection of sulfadimidine in water and dairy products by means of an evanescent wave optical biosensor

During recent years, there has been an increasing demand for the development of rapid biosensing technologies that could be performed outside the laboratory, for example on farms, near rivers, in food collection stations and store houses or in food production plants. Therefore, cost-effective and automatic detection methods are promising for onsite residual analysis in food and environmental monitoring. In this work, we propose an automatic, rapid, highly sensitive and reusable planar waveguide evanescent wave immunosensor (PWEI) for onsite determination of sulfadimidine (SM2) in water and dairy products. The PWEI is based on an indirect inhibition immunoassay that takes place at an optical transducer chip chemically modified with an analyte derivative. Fluorescence produced by labeled antibodies bound to the transducer is excited by the evanescent wave formed on the transducer surface and detected by photodiodes through a lock-in amplifier, which is inversely correlated with the analyte concentration. Each test cycle is completed automatically in 15 min. The optical transducer chip of the PWEI modified with the analyte derivative is robust and features high reusability, which allows for regeneration over 300 times without sensitivity loss. Under the optimized conditions, the dose–response curve established for SM2 shows a low detection limit of 0.06 μg L−1. The 50% inhibition concentration is 1.39 ± 0.08 μg L−1 with a linear working range from 0.19 μg L−1 to 10.10 μg L−1. The cross-reactivity towards organic compounds structurally similar to SM2 is negligible. The recoveries of SM2 in a variety of dairy products and natural water range from 80% to 107%. The PWEI features portable dimensions of 42 cm × 50 cm × 24 cm (length × width × height) and shows great prospects for the onsite measurement of SM2, when used in combination with the appropriate pretreatment.

TriPleX™ waveguide-based fluorescence biosensor for multichannel environmental contaminants detection

In order to realize the multi-analyte assays for environmental contaminants, an optical biosensor utilizing laser-induced fluorescence-based detection via the binding of biomolecules to the surface of an integrated TriPleX™ waveguide chip on a glass substrate (fused silica, FS) is described. As far as we know, this is the first demonstration of using the TriPleX™ technology to fabricate the waveguide chip on a FS substrate. The sensor consists of 32 individually addressable sensor patches, which were formed on the chip surface by exploiting 3 Y-junction splitters, creating four equal rows of eight evanescently excited windows in parallel. The basic low-loss SiO2/Si3N4 TriPleX™ waveguide configuration in combination with on-chip spotsize convertors allows for both high fiber-to-chip coupling efficiency and enables at the same time individually optimized high chip surface intensity and low patch-to-patch deviation. Moreover, the complementary metal-oxide-semiconductor compatible fabrication of waveguide chip allows for its mass production at low cost. By taking MC-LR, 2,4-D, atrazine and BPA as the model analytes, the as-proposed waveguide based biosensor was proven sensitive with the detection limits of 0.22 μg/L for MC-LR, 1.18 μg/L for 2, 4-D, 0.2 μg/L for atrazine and 0.06 μg/L for BPA. Recoveries of the biosensor towards simultaneous detection of MC-LR, 2, 4-D, atrazine and BPA in spiked real water samples varied from 84% to 120%, indicating the satisfactory accuracy of the established technology.