Determination of Lincomycin in Milk Using Cu-Based Metal-Organic Framework Adsorbent and Liquid Chromatography-Tandem Mass Spectrometry.

Antibiotic drug residues can adversely affect the human body. Lincomycin is a common veterinary drug that can form residues in foods of animal origin. However, the detection of trace residue levels of lincomycin residues in real samples is challenging. Here, a simple solid phase extraction (SPE) method was developed for the enrichment of lincomycin from cow milk samples before its detection by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The adsorbent used in the SPE was a Cu-based metal-organic framework (Cu-MOF) prepared by the solvothermal synthesis approach. The prepared MOFs were characterized using scanning electron microscopy (SEM), Fourier-transform infrared spectroscopy (FT-IR), X-ray diffractometry (XRD), differential thermogravimetric analysis (TG-DTA), and N2 adsorption-desorption experiments. The adsorption capacity (adsorption equilibrium, extraction time, pH), and elution solvent parameters were investigated. Under the optimized conditions of the HPLC-MS/MS method, lincomycin was detected in the linear range of 10-200 g/L with a detection limit of 0.013 ng/mL. Commercial milk samples were spiked with lincomycin, and a recovery rate between 92.3% and 97.2% was achieved. Therefore, the current method can be successfully applied for the enrichment and determination of lincomycin from milk samples.

Detection of ochratoxin A by fluorescence sensing based on mesoporous materials.

We developed a new ochratoxin A (OTA) aptamer biosensor to promptly detect OTA in food. Mesoporous silica nanoparticles were used as carriers, and aptamers were used as recognition probes and gating molecules. The fluorescent dye rhodamine 6G was loaded into mesoporous silica, and through electrostatic contact, the OTA aptamer was adsorbed on amino-modified mesoporous silica. The fluorescent dye released from the mesopore in the presence of OTA because of the conformational change induced in the aptamer by the target. The amount of ochratoxin was determined by measuring the fluorescence intensity. Our findings revealed a positive relationship between the fluorescence intensity and OTA concentration, with a limit of detection of 0.28 ng mL-1, and the detection range was 0.05-200 ng mL-1. The recovery rate was 80.7%-110.8% in real samples. The proposed approach is suitable for the quantification of other toxins.

Ultrasensitive Detection of 17ß-Estradiol (E2) Based on Multistep Isothermal Amplification.

17ß-Estradiol (E2) can cause an adverse effect on the human endocrine system even at the nanomolar level. Measurements of very low levels of E2 remain a critical challenge due to insufficient sensitivity. In this study, a multistep isothermal amplification fluorescence strategy was constructed, which could realize the exponential amplification of target E2. Specifically, strand displacement reaction (SDA), rolling circle amplification (RCA), and multiprimed rolling circle amplification (MRCA) were combined in a series to quantify trace complementary strand of E2 (cDNA). The E2 aptamer and cDNA were hybridized and modified on the magnetic beads. E2 could bind to its aptamer and cause the release of the cDNA. Then, cDNA would combine with the template DNA, initiating the SDA-RCA-MRCA. The molecular beacons, possessing low background signal, whose fluorescence was quenched in the state of chain folding, could be specifically recognized by the long single-stranded DNA (L-ssDNA) generated by the multistep isothermal amplification triggered by cDNA, and then the fluorescence of the molecular beacons could be restored. Therefore, the E2 could be quantitatively detected by the recovery fluorescence intensity. The fluorescence value showed a good linear relationship with the concentration of E2 in the range of 0.001836-183.6 nM, and the limit of detection (LOD) was as low as 63.09 fM. In addition, the recovery rates of this method spiked in milk and water were 80.8-107.0%, respectively. This method has the advantage of multistep isothermal amplification to obtain abundant fluorescence signals, which may provide a new possibility for highly sensitive detection of other small-molecule targets.

Dual Sensitization Smartphone Colorimetric Strategy Based on RCA Coils Gathering Au Tetrahedra and Its Application in the Detection of CK-MB.

In recent years, the combination of DNA nanotechnology and biosensing has been extensively reported. Herein, we attempted to develop a dual sensitization smartphone colorimetric strategy based on rolling circle amplification (RCA) coils gathering Au tetrahedra and explore its application. The dual sensitization effect of this strategy was achieved by rolling circle amplification (RCA) and Au tetrahedra. Under the initiation of the complementary DNA, a large number of ssDNA were generated, achieving amplification of the reaction signal. At the same time, due to the formation of Au tetrahedra, more gold nanoparticles could be gathered under the same conditions, and the signal would be amplified again. Using software ImageJ, the gray value of the reaction solution can be analyzed, detecting the target timely under the practical conditions of lack of equipment. By selecting aptamers with strong binding affinity, we applied this strategy to detect creatine kinase isoenzymes (CK-MB), showing a limit of detection of 0.8 pM, which performed well in actual detection and can meet the needs for real-time detection of CK-MB. Therefore, a universal detection platform was developed, which has broad application prospects in biosensing, clinical diagnosis, food detection, and other fields.

The orphan nuclear receptor Nur77 plays a vital role in BPA-induced PC12 cell apoptosis.

Bisphenol A (BPA) is a typical environmental endocrine disruptor that can migrate into organisms through skin contact, breathing, diet and various other approaches. The reproductive toxicity and neurotoxicity of BPA has been confirmed by several toxicological studies. However, the neurotoxicity of BPA is still controversial. In the present study, we used PC12 cells as a model to investigate the mechanism of BPA-induced neuronal apoptosis. BPA exposure reduced cell viability, altered cell morphology and aggravated intracellular Lactate dehydrogenase (LDH) release, intracellular Ca2+ concentration, Reactive oxygen species (ROS) levels, apoptosis and the reduction in the mitochondrial transmembrane potential (ΔΨm). Moreover, the results of the Western blot (WB) and Real-time quantitative polymerase chain reaction (RT-qPCR) assays indicated that the expression levels of Nur77 in the BPA group were down-regulated and accompanied by the downregulation of the NF-κb/Bcl-2 proteins and the upregulation of cleaved-caspase 3, which is a marker of apoptosis. However, these changes were significantly reversed with the upregulation of the Nur77 protein by introducing plasmids carrying the nur77 gene. These results indicated that BPA-induced apoptosis was closely related to Nur77-mediated inhibition of the NF-κb/Bcl-2 pathway.

Dual-emission ratio fluorescent probes based on carbon dots and gold nanoclusters for visual and fluorescent detection of copper ions.

Blue carbon dots (BCDs) and red gold nanoclusters modified by bovine serum albumin (BSA-Au NCs) were selected as luminescent nanomaterials, and the nanohybrid materials were successfully prepared and applied to the fluorescent measurement of copper ions. The prepared ratio fluorescent probe has two emission peaks near 452 and 654 nm under an excitation of 330 nm. The fluorescence intensity was gradually quenched because Au NCs was aggregated in the presence of Cu2+, resulting in a gradation of the fluorescent color from red to pink to purple to blue for visual detection. BCDs have almost the same fluorescence intensity due to their light stability and chemical inertness to Cu2+ and serve as a background reference in the sensing system. Under the optimal condition, the detection limit (LOD) is 16 nM, the linear range is 0.05-1.85 µM, and the coefficient of determination R2 is 0.9987 for copper determination. Compared with other single probes, the ratio fluorescent probe in the current study has good water solubility, low cytotoxicity, and is easy to synthesize. The nanoprobe provides a high-quality and sensitive visible light platform for monitoring copper ions. Graphical abstract.

A low-field nuclear magnetic resonance DNA-hydrogel nanoprobe for bisphenol A determination in drinking water.

A low-field nuclear magnetic resonance (LF-NMR) DNA-hydrogel (LNDH) nanoprobe was designed for bisphenol A (BPA) determination. It consists of Fe3O4 superparamagnetic iron oxide nanoparticles (SPIONs) and a DNA-hydrogel technology. Fe3O4 SPIONs were encapsulated in the DNA-hydrogel to form an aggregated state. After adding BPA, the gel system transformed into a sol gel due to the target-aptamer specific binding. The coated gathered particles dispersed and thus, the relaxation time T2 declined. The LNDH nanoprobe was developed to realize a simple, sensitive, and effective BPA determination method without repeated magnetic separation steps. Under the optimal experimental conditions, the determination range of the LNDH biosensor was 10-2~102 ng mL-1 and the limit of determination was 0.07 ng mL-1. The LNDH nanoprobe was applied to two kinds of water samples (tap water and bottled water). The recovery ranged from 87.85 to approximately 97.87%. This strategy offered a new method to detect BPA by LF-NMR. It is also expected to be applicable in related fields of food safety determination, environmental monitoring, and clinical diagnosis. Graphical abstract Schematic presentation of LNDH biosensor. Acrydite-modified ssDNA was copolymerized with acrylamide to form linear conjugates PS-A/B, adding aptamer and SPIONs to form DNA-hydrogel. When aptamer captured the target, the hydrogel was destroyed to disperse the coated SPIONs. T2 relaxation time declined.

An ultrasensitive sensor based on quantitatively modified upconversion particles for trace bisphenol A detection.

Bisphenol A (BPA) is one of the endocrine-disrupting chemicals which might cause reproductive and endocrine system diseases, and poses a serious threat to the ecosystem and human health. This paper reports an ultrasensitive sensor for trace BPA detection employing fluorescence resonance energy transfer (FRET) between modified upconversion nanoparticles (UCNPs) and tetramethylrhodamine. To circumvent the problems of low luminous efficiency of FRET and low sensitivity of sensor, the upconversion nanoparticles with very strong fluorescence efficiency were prepared and quantitatively modified. Results showed that the concentrations of amino groups and streptavidin were 43 nmol/mg and 6.12 µg/mg on the surface of the UCNPs, respectively. Under the optimal detection conditions, the peak intensity of UCNPs at 547 nm was linear with the logarithm of the BPA concentration with the detection limit of 0.05 ng/mL. Without complicated pre-processing, the recoveries were in general between 91.0 and 115.0% in tap water, river water, and disposable paper cup water. Therefore, the proposed sensor is suitable for effective sensing of trace BPA in water samples. Graphical abstract ᅟ.

An aptamer-based fluorometric zearalenone assay using a lighting-up silver nanocluster probe and catalyzed by a hairpin assembly.

An enzyme-free fluorometric assay is described for the determination of zearalenone (ZEN). The method combines (a) catalyzed hairpin assembly (CHA), (b) ultrahigh fluorescent light-up G-rich DNA sequences in proximity to silver nanoclusters (Ag NCs), and (c) the use of aptamers (Apt). In the presence of ZEN, the inhibit sequence (Inh) is released from the aptamer-trigger sequence (Apt-T) via the binding of ZEN and the aptamer of Apt-T. The free Apt-T acts as a switch that opens the hairpins H1 and H2 to generate H1-H2 complex. The released Apt-T is available to trigger the next round of CHA between H1 and H2. Finally, the hybridization between H1 and the Ag NCs probe (P) causes the G-rich sequence to be in close proximity to the dark Ag NCs encapsulated by P. This leads to highly efficient lighting up of the Ag NCs and the production of amplified fluorescence with excitation/emission peaks at 575/628 nm. Under the optimized conditions, a linear correlation was observed with concentrations ranging from 1.3 pg mL-1 to 100 ng mL-1, and the limit of detection was 0.32 pg mL-1 (at S/N = 3). The method was successfully validated by analyzing maize and beer for levels of ZEN after a simple sample preparation procedure. Graphical abstractSchematic of the assay. The inhibit sequence (Inh) is released from aptamer-trigger sequence (Apt-T) via binding of ZEN and aptamer. The free Apt-T triggers catalyzed hairpin assembly (CHA).G-rich DNA is in proximity to silver nanoclusters (Ag NCs) and fluorescence intensity increases to detect ZEN.

Competitive fluorometric assay for the food toxin T-2 by using DNA-modified silver nanoclusters, aptamer-modified magnetic beads, and exponential isothermal amplification.

The authors describe an aptamer based assay for the mycotoxin T-2. The method is making use of exponential isothermal amplification reaction (EXPAR) and fluorescent silver nanoclusters (AgNCs). Free T-2 and cDNA (which is a DNA that is partially complementary to the aptamer) compete for binding to aptamer-modified magnetic beads. The cDNA collected by magnetic separation can be used as a primer to trigger EXPAR to obtain ssDNA. The C-base-rich ssDNA binds and reduces Ag(I) ion to form fluorescent AgNCs. Fluorescence is measured at excitation/emission wavelengths of 480/525 nm. T-2 can be detected by fluorometry with a detection limit as low as 30 fg·mL-1. The method was applied to analyse spiked oat and corn, and the average recoveries ranged from 97.3 to 102.3% and from 95.9 to 107.5%, respectively. The results were in good agreement with data of the commercial ELISA kit. The assay is highly sensitive, has a wide analytical range, good specificity and reliable operation. It provides a promising alternative for the standard method for quantitative detection of T-2. Graphical abstract Schematic presentation of fluorometric assay for T-2 based on aptamer-functionalized magnetic beads exponential, isothermal amplification reaction (EXPAR) and fluorescent silver nanoclusters (AgNCs).

Upconversion fluorescent aptasensor for bisphenol A and 17ß-estradiol based on a nanohybrid composed of black phosphorus and gold, and making use of signal amplification via DNA tetrahedrons.

This study describes an upconversion fluorescent aptasensor based on black phosphorus nanohybrids and self-assembled DNA tetrahedrons dual-amplification strategy for rapid detection of the environmental estrogens bisphenol A (BPA) and 17ß-estradiol (E2). Tetrahedron complementary DNAs (T-cDNAs) were self-assembled in an oriented fashion on a 2D nanohybrid composed of black phosphorus (BP) and gold to give a materials of architecture BP-Au@T-cDNAs. In parallel, core-shell upconversion nanoparticles were modified with aptamers (UCNPs@apts) and used as capture probes. On complementary pairing, the BP-Au@T-cDNA quench the fluorescence of UCNPs@apts (measured at an excitation wavelength 808 nm and at main emission peaks at 545 nm and 805 nm.) Compared with single-stranded probes based on black phosphorus and gold, the dual-amplification strategy increases quenching efficiency by nearly 25%-30% and reduces capture time to 10 min. This is due to the higher optical absorption of 2D nanohybrid and the reduction of steric hindrance by T-cDNAs. Exposure to BPA or E2 cause the release of UCNPs@apts from the BP-Au@T-cDNAs due to stronger binding between aptamer and analyte. Hence, fluorescence recovers at 545 nm for BPA and 805 nm for E2. Based on these findings, a dually amplified aptamer assay was constructed that covers the 0.01 to 100 ng mL-1 BPA concentration range, and the 0.1 to 100 ng mL-1 E2 concentration range. The detection limits are 7.8 pg mL-1 and 92 pg mL-1, respectively. This method was applied to the simultaneous determination of BPA and E2 in spiked samples of water, food, serum and urine. Graphical abstract Schematic presentation of novel quenching probes designed by tetrahedron complementary DNAs oriented self-assembled on the surface of black phosphorus/gold nanohybrids. Combined with aptamer-modified upconversion nanoparticles, a dual-amplification self-assembled fluorescence nanoprobe was constructed for simultaneous detection of BPA and E2.

Correction to: An aptamer-based fluorometric zearalenone assay using a lighting-up silver nanocluster probe and catalyzed by a hairpin assembly.

The published version of this article, unfortunately, contains an error regarding authors for correspondence.

Upconversion Fluorescent Aptasensor for Polychlorinated Biphenyls Detection Based on Nicking Endonuclease and Hybridization Chain Reaction Dual-Amplification Strategy.

A novel upconversion fluorescent aptasensor based on hybridization chain reaction and nicking endonuclease has been developed for detection of polychlorinated biphenyls (PCBs). It combined the dual advantages of UCNPs and HCR. Two harpins (H1 and H2) were first designed according to the partial complementary sequence (cDNA) of the PCB72/106. Since the aptamer specifically recognized the target, the cDNA was detached from the magnetic microspheres (MMPs). The cDNA could initiate hybridization chain reaction (HCR) and open the stems of H1 and H2. After the addition of nicking endonuclease, UCNPs were further away from the quenchers (BHQ-1). Hence, the fluorescence intensity of upconversion nanoparticals (UCNPs) could be restored via fluorescence resonance energy transfer (FRET). Therefore, the fluorescence of UCNPs was directly proportional to concentration of PCB72/106, which was the basis for the quantification of PCB72/106. PCB72/106 could be analyzed within the ranges of 0.004 to 800 ng/mL with a detection limit of 0.0035 ng/mL ( S/ N = 3). The aptasensor was also used for the detection of water and soil samples, and the average recoveries ranged from 93.4% to 109.7% and 83.2% to 118.5%, respectively. The relative standard deviations (RSDs) were all below 3.2%. The signal was first amplified through HCR and further amplified with the help of nicking endonuclease. This work also provided the opportunity to develop fluorescent aptasensors for other targets using this dual-amplification strategy.

Michael-Addition-Mediated Photonic Crystals Allow Pretreatment-Free and Label-Free Sensoring of Ciprofloxacin in Fish Farming Water.

Abuse of antibiotics results in a large number of antibiotics residues in the environment and even causes the problem of "super bacteria". Therefore, it is crucial to develop a powerful analytic method to monitor antibiotics quickly and simply. Photonic crystal (PC), as a sensing material, has promising application prospects. Herein, we try to use PC to realize pretreatment-free and label-free detection of Ciprofloxacin (CF) through Michael addition reaction. The recognition process is carried out by the Michael addition reaction between the piperazine group of CF and the o-benzoquinone group on the PC. The monodisperse microspheres with o-benzoquinone groups are prepared by polymerization and oxidation and then stacked to form PC. During the detection, the peak intensity of the PC decreases with the increasing CF concentration, and the linear range is from 2 to 512 µg/L. The limit of detection (LOD) is 0.76 µg/L. Furthermore, the PC retains 97% of the initial response after storage in a Petri dish at room temperature for 1 month, which shows that it has good stability. Moreover, CF in fish farming water can be detected directly without any pretreatment and label, and the results are in good accordance with the LC-MS-MS results. This Michael-addition-mediated PC is accurate, easily prepared, cost-efficient, and long-term stable. In addition, it is environmentally friendly, because little organic solvent is needed during both the preparation and the detection.

Turn-on fluorometric immunosensor for diethylstilbestrol based on the use of air-stable polydopamine-functionalized black phosphorus and upconversion nanoparticles.

The preparation of air-stable black phosphorus (BP) is challenging because atomic layers of BP degrade rapidly on exposure to oxygen. A strategy is presented for the synthesis of BP functionalized with polydopamine (PDA/BP). Dopamine was self-polymerized to yield polydopamine (PDA) which then was used to coat the surface of BP. PDA can be easily reduced and this prevents BP degradation. PDA/BP also is a viable matrix for the adsorption of proteins due to the presence of functional groups. Without any chemical activation, diethylstilbestrol (DES)-specific monoclonal antibody was adsorbed on the PDA/BP surface. PDA/BP quenches the fluorescence antigen-modified NaYF4:Yb,Ho,Nd upconversion nanoparticles (UCNPs; photoexcited at 808 nm) via specific immuno recognition. Exposure to DES causes the dissociation of UCNP from the PDA/BP surface and fluorescence at 475, 525, 545 and 660 nm to recover. This is due to the DES competition with antigen for binding to the antibody. Based on this competitive immuno mechanism, a turn-on fluorometric immunoassay was constructed. It has a response that covers the 0.1 to 1000 ng mL-1 DES concentration range with a detection limit of 83 pg mL-1. This method was successfully applied to the determination of DES in spiked food and human urine samples. Graphical abstract Air-stable polydopamine-functionalized black phosphorus was obtained by modification of black phosphorus with polydopamine and then was coupled with specific monoclonal antibody. Combined with antigen-modified upconversion nanoparticles, a turn-on fluorometric immunoassay was constructed to detect diethylstilbestrol.

[Rapid detection of diethylstilbestrol using a quartz crystal microbalance with gold nanoparticals amplification].

OBJECTIVE: To develop a quartz crystal microbalance (QCM) immunosensor with high sensitivity and selectivity for the rapid detection of diethylstilbestrol. METHODS: Dextran was used as reducing agent for preparing gold nanoparticles (AuNPs) with the size of 40 nm. The AuNPs were coupled with anti-DES antibody after amination. A monolayer was generated after immersing the quartz crystal into the solution of 5 mmol/L 11-mercaptoundecanoic acid(MUA) for 16 hours. After the monolayer was activated by 1-ethyl-3-(3-dimethylaminopropry) carbodiimide hydrochloride (EDC·HCl) and N-hydrosuccinimide (NHS), 20 µl of 2.2 mg/ml DES-HS-BSA was dropped onto the surface of crystal to prepare a sensitive membrane which can recognize DES specifically. Then, 50 µl of 1 mol/L ethanolamine (pH 8.5) was used to seal the carboxylic groups to make the sensitive membrane which could identify DES specifically. QCM immunosensor was used as detection platform to optimize the reaction conditions. Under the optimized conditions, 10 µl of 28 µg/ml AuNPs-antibody was mixed with 10 µl of 0.03-2.5 µg/ml DES, and the mixture was added on the sensitive membrane. QCM immunosensor was used to detect the signals and the standard curve was obtained at the same time. The detection limit was calculated based on the standard curve. The specificity was evaluated by testing DES and its analogues with the same concentration. RESULTS: The optimized concentration for the immobilization of DES-HS-BSA on the surface of QCM was 2.2 mg/ml. The optimized concentration for coupling anti-DES antibody with AuNPs was 7 µg/ml and 15 nmol/L, respectively. The optimized concentration of AuNPs-antibody was 14 µg/ml. The logarithm of DES concentration was proportional to the frequency shift in the range of 0.16-500 ng/ml, Δf=-24.170 lgCDES+69.71, R(2)=0.998. The detection limit of this method was 0.13 ng/ml. DES analogues could not influence the detection of DES obviously, so the sensor had good specificity. CONCLUSION: The quartz crystal microbalance immunosensor with gold nanoparticals amplification could detect DES sensitively and rapidly.

Selection of bisphenol A - single-chain antibodies from a non-immunized mouse library by ribosome display.

Developing reagents with high affinity and specificity are critical to detect the environmental hormones or toxicants. Ribosome display technology has been widely used in functional protein or peptide screening and in directed evolution of protein molecules in vitro. In this study, single-chain variable fragments (scFvs) against bisphenol A (BPA) were selected from a library constructed from splenocytes of non-immunized mice. After five rounds of selection, the selected scFvs bound to BPA with high affinity. Indirect competitive enzyme-linked immunosorbent assay (ELISA) was introduced to screen the antibody affinity and specificity to BPA. The equilibrium dissociation constants (KDS) of one clone was 1.76µM as determined by surface plasmon resonance (SPR). This study indicated that ribosome display can isolate binders to small molecules from a non-immunized naive library without any in vivo steps and can generate recombinant antibodies efficiently and rapidly. In addition, this study provides a methodological framework for detection of small molecules using recombinant antibodies.

A core-shell-structured molecularly imprinted polymer on upconverting nanoparticles for selective and sensitive fluorescence sensing of sulfamethazine.

A core-shell structured molecularly imprinted polymer on upconverting nanoparticles (UCNPs@MIP) was synthesized for the fluorescence (FL) sensing of sulfamethazine (SMZ). Hexagonal UCNPs were synthesized by the solvothermal method, then coated with a thin silica shell and modified with vinyl groups. Finally, surface polymerization was initiated among the vinyl groups, the functional monomers and cross-linking agents by the initiator. The MIP synthesized by this procedure was anchored on the surface of UCNPs, possessed better site accessibility and lower transfer resistance for the target molecule compared to bulk imprinted materials. The obtained UCNPs@MIP showed good binding capacity, fast response, high selectivity and specificity to the SMZ. The FL intensity of the UCNPs@MIP decreased sensitively with the increasing concentration of SMZ in the range of 50-700 ng mL(-1), the detection limit was 34 ng mL(-1) (S/N = 3). The UCNPs@MIP was successfully applied to the detection of SMZ in chicken samples. Thanks to the unique near-infrared (NIR) excitation nature of UCNPs, the chicken meat only needed some simple extraction procedures before FL detection, no complex purifications were required. The average recoveries ranged from 96.01% to 98.90%, with relative standard deviations (RSDs) below 4.5%. This work offers a novel sensing system that combined the advantages of upconverting nanotechnology and molecularly imprinted technology.

Sensitive detection of atrazine in tap water using TELISA.

A highly sensitive flow injection analysis (FIA)-based thermal enzyme-linked immunoassay, TELISA, was developed for the rapid detection of atrazine (ATZ) in tap water. ATZ and ß-lactamase-labeled ATZ were employed in a competitive immunoassay using a monoclonal antibody (mAb). After the off-column liquid-phase competition, the mAb was captured on the Protein G Sepharose™ 4 Fast Flow (PGSFF) column support material. Injected ß-lactamase substrate ampicillin was degraded by the column-bound ATZ-ß-lactamase, generating a detectable heat signal. Several assay parameters were optimized, including substrate concentration, flow rates and regeneration conditions, as well as the mAb and ATZ-ß dilution ratios and concentrations. The assay linear range was 0.73-4.83 ng mL(-1) with a detection limit of 0.66 ng mL(-1). An entire heat signal requires 10 min for generation, and the cycle time is less than 40 min. The results were reproducible and stable. ATZ-spiked tap water samples exhibited a recovery rate of 103%-116%, which correlated with the UHPLC-MS/MS measurements. We attributed this significant increase in sensitivity over our previously published work to the following factors: (i) the capture of already-formed immune complexes on the column via immobilized Protein G, which eliminated chemical immobilization of the antibody; (ii) off-column preincubation allows the formation of immune complexes under nearly ideal conditions; and (iii) multiple buffers can be used to, in one case, enhance immune-complex formation and in the other to maximize enzymatic activity. Furthermore, the scheme creates a universal assay platform in which sensing is performed in the off-column incubation and detection after capture in the enzyme thermistor (ET) detector, which opens up the possibility of detecting any antigen for which antibodies were available.

A novel opal closest-packing photonic crystal for naked-eye glucose detection.

A novel opal closest-packing (OCP) photonic crystal (PC) is successfully prepared for naked-eye glucose detection. This PC is fabricated via a vertical convective self-assembly method with a new type of monodisperse microsphere polymerized by co-monomers, namely, methyl methacrylate (MMA), N-isopropylacrylamide (NIPA), and 3-acrylamidophenylboronic acid (AAPBA). The OCP PC has high stability and periodically-ordered structure, showing the desired structural color. The proposed PC material displays a red shift and reduced reflection intensity when detecting glucose molecules. The red-shift wavelength reaches 75 nm, which clearly changes the structural color from brilliant blue to emerald green. This visually distinguishable color change facilitates the detection of the glucose concentrations from 3 to 20 mm, which demonstrates the potential of the opal PC material for naked-eye detection. Thus, the novel PMMA­NIPA­AAPBA OCP PC is a simply prepared and sensitive material, which shows promising use in the diagnosis of diabetes mellitus and in real-time monitoring of diabetes. Different types of appropriated recognition groups are expected to be introduced into the 3D OCP PC to form new functional materials or chemical sensors, which will extensively broaden the PC material application.