Light-addressable potentiometric sensors (LAPS) are of great interest in bioimaging applications such as the monitoring of concentrations in microfluidic channels or the investigation of metabolic and signaling events in living cells. By measuring the photocurrents at electrolyte-insulator-semiconductor (EIS) and electrolyte-semiconductor structures, LAPS can produce spatiotemporal images of chemical or biological analytes, electrical potentials and impedance. However, its commercial applications are often restricted by their limited AC photocurrents and resolution of LAPS images. Herein, for the first time, the use of 1D semiconducting oxides in the form of ZnO nanorods for LAPS imaging is explored to solve this issue. A significantly increased AC photocurrent with enhanced image resolution has been achieved based on ZnO nanorods, with a photocurrent of 45.7 ± 0.1 nA at a light intensity of 0.05 mW, a lateral resolution as low as 3.0 μm as demonstrated by images of a PMMA dot on ZnO nanorods and a pH sensitivity of 53 mV/pH. The suitability of the device for bioanalysis and bioimaging was demonstrated by monitoring the degradation of a thin poly(ester amide) film with the enzyme α-chymotrypsin using LAPS. This simple and robust route to fabricate LAPS substrates with excellent performance would provide tremendous opportunities for bioimaging.
Drug combinations are commonly used to treat various diseases to achieve synergistic therapeutic effects or to alleviate drug resistance. Nevertheless, some drug combinations might lead to adverse effects, and thus, it is crucial to explore the mechanisms of drug interactions before clinical treatment. Generally, drug interactions have been studied using nonclinical pharmacokinetics, toxicology, and pharmacology. Here, we propose a complementary strategy based on metabolomics, which we call interaction metabolite set enrichment analysis, or iMSEA, to decipher drug interactions. First, a digraph-based heterogeneous network model was constructed to model the biological metabolic network based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Second, treatment-specific influences on all detected metabolites were calculated and propagated across the whole network model. Third, pathway activity was defined and enriched to quantify the influence of each treatment on the predefined functional metabolite sets, i.e., metabolic pathways. Finally, drug interactions were identified by comparing the pathway activity enriched by the drug combination treatments and the single drug treatments. A data set consisting of hepatocellular carcinoma (HCC) cells that were treated with oxaliplatin (OXA) and/or vitamin C (VC) was used to illustrate the effectiveness of the iMSEA strategy for evaluation of drug interactions. Performance evaluation using synthetic noise data was also performed to evaluate sensitivities and parameter settings for the iMSEA strategy. The iMSEA strategy highlighted synergistic effects of combined OXA and VC treatments including the alterations in the glycerophospholipid metabolism pathway and glycine, serine, and threonine metabolism pathway. This work provides an alternative method to reveal the mechanisms of drug combinations from the viewpoint of metabolomics.
Rayleigh surface acoustic waves (SAWs) have been demonstrated as a powerful and effective means for driving a wide range of microfluidic actuation processes. Traditionally, SAWs have been generated on piezoelectric substrates, although the cost of the material and the electrode deposition process makes them less amenable as low-cost and disposable components. As such, a "razor-and-blades" model that couples the acoustic energy of the SAW on the piezoelectric substrate through a fluid coupling layer and into a low-cost and, hence, disposable silicon superstrate on which various microfluidic processes can be conducted has been proposed. Nevertheless, it was shown that only bulk vibration in the form of Lamb waves can be excited in the superstrate, which is considerably less efficient and flexible in terms of microfluidic functionality compared to its surface counterpart, that is, the SAW. Here, we reveal an extremely simple way that quite unexpectedly and rather nonintuitively allows SAWs to be generated on the superstrate-by coating the superstrate with a thin gold layer. In addition to verifying the existence of the SAW on the coated superstrate, we carry out finite-difference time domain numerical simulations that not only confirm the experimental observations but also facilitate an understanding of the surprising difference that the coating makes. Finally, we elucidate the various power-dependent particle concentration phenomena that can be carried out in a sessile droplet atop the superstrate and show the possibility for simply carrying out rapid and effective microcentrifugation-a process that is considerably more difficult with Lamb wave excitation on the superstrate.
We present a compact, cost-effective, label-free, real-time biosensor based on long-range surface plasmon polariton (LRSPP) gold (Au) waveguides for the detection of dengue-specific immunoglobulin M (IgM) antibody, and we demonstrate detection in actual patient blood plasma samples. Two surface functionalization approaches are proposed and demonstrated: a dengue virus serotype 2 (DENV-2) functionalized surface to capture dengue-specific IgM antibody in blood plasma and the reverse, a blood plasma functionalized surface to capture DENV-2. The results obtained via these two surface functionalization approaches are comparable to, or of greater quality, than those collected by conventional IgM antibody capture enzyme linked immunosorbent assay (MAC-ELISA). Our second functionalization approach was found to minimize nonspecific binding, thus improving the sensitivity and accuracy of the test. We also demonstrate reuse of the biosensors by regenerating the sensing surface down to the virus (or antibody) level or down to the bare Au.
A novel sequential three-dimensional gas chromatography-high-resolution time-of-flight mass spectrometry (3D GC-accTOFMS) approach for profiling secondary metabolites in complex plant extracts is described. This integrated system incorporates a nonpolar first-dimension (1Dnp) separation step, prior to a microfluidic heart-cut (H/C) of a targeted region(s) to a cryogenic trapping device, directly followed by the rapid reinjection of a trapped solute into a polar second-dimension (2DPEG) column for multidimensional separation (GCnp-GCPEG). For additional separation, the effluent from 2DPEG can then be modulated according to a comprehensive 2D GC process (GC×GC), using an ionic liquid phase as a third-dimension (3DIL) column, to produce a sequential GCnp-GCPEG×GCIL separation. Thus, the unresolved or poorly resolved components, or regions that require further separation, can be precisely selected and rapidly transferred for additional separation on 2D or 3D columns, based on the greater separation realized by these steps. The described integrated system can be used in a number of modes, but one useful approach is to target specific classes of compounds for improved resolution. This is demonstrated through the separation and detection of the oxygenated sesquiterpenes in hop ( Humulus lupulus L.) essential oil and agarwood ( Aquilaria malaccensis) oleoresin. Improved resolution and peak capacity were illustrated through the progressive comparison of the tentatively identified components for GCnp-GCPEG and GCnp-GCPEG×GCIL methods. Relative standard deviations of intraday retentions (1 tR, 2 tR,, and 3 tR) and peak areas of ≤0.01, 0.07, 0.71, and 7.5% were achieved. This analytical approach comprising three GC column selectivities, hyphenated with high-resolution TOFMS detection, should be a valuable adjunct for the improved characterization of complex plant samples, particularly in the area of plant metabolomics.
A general purpose enzyme-based amperometric electrochemical genosensor assay was developed wherein polymerase chain reaction (PCR) amplicons labeled with both biotin and fluorescein were detected with peroxidase-conjugated antifluorescein antibody on a screen-printed carbon electrode (SPCE). As a proof of principle, the response selectivity of the genosensor was evaluated using PCR amplicons derived from lolB gene of Vibrio cholerae. Factors affecting immobilization, hybridization, and nonspecific binding were optimized to maximize sensitivity and reduce assay time. On the basis of the background amperometry signals obtained from nonspecific organisms and positive signals obtained from known V. cholerae, a threshold point of 4.20 microA signal was determined as positive. Under the optimum conditions, the limit of detection (LOD) of the assay was 10 CFU/mL of V. cholerae. The overall precision of this assay was good, with the coefficient of variation (CV) being 3.7% using SPCE and intermittent pulse amperometry (IPA) as an electrochemical technique. The assay is sensitive, safe, and cost-effective when compared to conventional agarose gel electrophoresis, real-time PCR, and other enzyme-linked assays for the detection of PCR amplicons. Furthermore, the use of a hand-held portable reader makes it suitable for use in the field.
2D nanosheets (NSs) have been widely used in drug-related applications. However, a comprehensive investigation into the cytotoxicity mechanism linked to the redox activity is lacking. In this study, with cytochrome c (Cyt c) as the model biospecies, the cytotoxicity of 2D NSs was evaluated systematically based on their redox effect with microfluidic techniques. The interface interaction, dissolution, and redox effect of 2D NSs on Cyt c were monitored with pulsed streaming potential (SP) measurement and capillary electrophoresis (CE). The relationship between the redox activity of 2D NSs and the function of Cyt c was evaluated in vitro with Hela cells. The results indicated that the dissolution and redox activity of 2D NSs can be simultaneously monitored with CE under weak interface interactions and at low sample volumes. Both WS2 NSs and MoS2 NSs can reduce Cyt c without significant dissolution, with reduction rates measured at 6.24 × 10-5 M for WS2 NSs and 3.76 × 10-5 M for MoS2 NSs. Furthermore, exposure to 2D NSs exhibited heightened reducibility, which prompted more pronounced alterations associated with Cyt c dysfunction, encompassing ATP synthesis, modifications in mitochondrial membrane potential, and increased reactive oxygen species production. These observations suggest a positive correlation between the redox activity of 2D NSs and their redox toxicity in Hela cells. These findings provide valuable insight into the redox properties of 2D NSs regarding cytotoxicity and offer the possibility to modify the 2D NSs to reduce their redox toxicity for clinical applications.
DNAzymes have emerged as an important class of sensors for a wide variety of metal ions, with florescence DNAzyme sensors as the most widely used in different sensing and imaging applications because of their fast response time, high signal intensity, and high sensitivity. However, the requirements of an external excitation light source and its associated power increase the cost and size of the fluorometer, making it difficult to be used for portable detections. To overcome these limitations, we report herein a DNAzyme sensor that relies on chemiluminescence resonance energy transfer (CRET) without the need for external light. The sensor is constructed by combining the functional motifs from both Pb2+-dependent 8-17 DNAzyme conjugated to fluorescein (FAM) and hemin/G-quadruplex that mimics horseradish peroxidase to catalyze the oxidation of luminol by H2O2 to yield chemiluminescence. In the absence of Pb2+, the hybridization between the enzyme and substrate strands bring the FAM and hemin/G-quadruplex in close proximity, resulting in CRET. The presence of Pb2+ ions can drive the cleavage on the substrate strand, resulting in a sharp decrease in the melting temperature of hybridization and thus separation of the FAM from hemin/G-quadruplex. The liberated CRET pair causes a ratiometric increase in the donor's fluorescent signal and a decrease in the acceptor signal. Using this method, Pb2+ ions have been measured rapidly (<15 min) with a low limit of detection at 5 nM. By removing the requirement of exogenous light excitation, we have demonstrated a simple and portable detection using a smartphone, making the DNAzyme-CRET system suitable for field tests of lake water. Since DNAzymes selective for other metal ions or targets, such as bacteria, can be obtained using in vitro selection, the method reported here opens a new avenue for rapid, portable, and ratiometric detection of many targets in environmental monitoring, food safety, and medical diagnostics.