Multiplex digital microfluidics using serial controls and its applications in glucose sensing.

Digital microfluidics (DMF) has found great applications in vitro diagnostics (IVD). Compared to the microfabrication-based DMF, printed circuit board (PCB)-based DMF is more economical and compatible with existing IVD instruments. Despite that, current PCB-based DMF is oftentimes limited by the available droplets that can be controlled simultaneously, compromising their throughput and applications as point-of-care tools. In this work, a platform that simultaneously controls multiple PCB-based DMF plates was constructed. The software and hardware were first developed, followed by the reliability tests. Colorimetric analysis of glucose was applied to the PCB-based DMF, demonstrating the capability of this platform. With the high throughput enabled by simultaneous operations of multiple plates, this PCB-based DMF can potentially allow point-of-care testing with low cost for resource-limited settings.

Ligustrazine induces viability, suppresses apoptosis and autophagy of retinal ganglion cells with ischemia/reperfusion injury through the PI3K/Akt/mTOR signaling pathway.

Ligustrazine, an alkaloid monomer extracted from Chuanxiong Rhizoma, has the function of protecting nerve cells. However, the effect and mechanism of ligustrazine on retinal ischemia/reperfusion (I/R) injury still need to be clarified. In our study, retinal ganglion cells (RGC-5) were used to establish a retinal I/R injury model by anaerobic cultivation. Cell viability, autophagy, and apoptosis were evaluated by cell counting kit 8 assay, transmission electron microscopy, and TUNEL staining after treatment with ligustrazine, PI3K inhibitor Ly294002, and/or mTOR inhibitor rapamycin, respectively. Besides, the levels of PI3K/Akt/mTOR pathway and autophagy-related proteins were determined by western blot. Moreover, one-way ANOVA was adopted for inter-group comparisons of measurement data. Our results demonstrated that low-concentration ligustrazine significantly enhanced cell viability and suppressed cell autophagy and apoptosis of RGC-5 cells after I/R injury, suggesting the protective effect of low-concentration ligustrazine on retinal I/R injury. Moreover, the alleviating effect of ligustrazine on RGC-5 with retinal I/R injury was mechanistically associated with the activation of the PI3K/Akt/mTOR pathway. In conclusion, low-concentration ligustrazine has a significant protective effect on RGC-5 cells with retinal I/R injury by activating the PI3K/Akt/mTOR pathway.

Ligustrazine protects against chronic hypertensive glaucoma in rats by inhibiting autophagy via the PI3K-Akt/mTOR pathway.

PURPOSE: Glaucoma is a leading cause of global irreversible blindness, and characterized by the progressive loss of retinal ganglion cells (RGCs). Ligustrazine (TMP) is a natural product that has shown beneficial effects on various diseases. This study aimed to determine whether ligustrazine produces a therapeutic effect on glaucoma and to investigate its underlying mechanisms. METHODS: A rat chronic hypertensive glaucoma model was induced by episcleral vein cauterization (EVC). Adult Sprague-Dawley (SD) rats were intraperitoneally administered TMP at a dose of 80 mg/kg once a day, from two days before EVC to one month after EVC. To elucidate the role of the mammalian target of rapamycin (mTOR) and phosphoinositide 3-kinase (PI3K), TMP-treated experimental rats were co-treated with the mTOR inhibitor rapamycin (5 mg/kg) or the PI3K inhibitor Ly294002 (10 mg/kg). The intraocular pressure (IOP) of the experimental and control rats was measured every six days. Retinal cells were examined by hematoxylin-eosin and terminal deoxynucleotidyltransferase-mediated biotinylated UTP nick end labeling (TUNEL) staining, as well as transmission electron microscopy. Immunohistochemistry and western blot analysis were performed to measure proteins involved in apoptosis and autophagy. RESULTS: Ligustrazine protected retinal cells from death in experimental glaucoma rats, which was not due to the lowering of IOP, but could be attributable to direct suppression of retinal cell apoptosis. In glaucoma rats, autophagy was markedly activated in retina cells, as evidenced by increased numbers of autophagosomes and the expression of autophagy-related proteins (ATG5 and LC3-II/I). Notably, such alterations in glaucoma rats were almost completely reversed by ligustrazine. The suppressive effects of ligustrazine on apoptosis and autophagy of retina cells were markedly attenuated by the mTOR inhibitor rapamycin or the PI3K inhibitor Ly294002. Additionally, ligustrazine significantly increased the protein levels of phosphorylated PI3K (p-PI3K), protein kinase B (p-Akt), and mTOR (p-mTOR) in glaucoma rats, whereas such increases were attenuated by rapamycin or Ly294002. CONCLUSIONS: These results demonstrate that ligustrazine is protective in experimental glaucoma by inhibiting autophagy via the activation of the PI3K-Akt/mTOR pathway, providing compelling evidence that ligustrazine is potentially therapeutic for patients with glaucoma.

Porous graphene-black phosphorus nanocomposite modified electrode for detection of leptin.

Leptin is a vital biomarker of non-alcoholic fatty liver (NAFLD), and its evaluation of the concentration level in vivo is of great significance to NAFLD diagnosis. Therefore, it is pressing to develop a method for rapid and sensitive detection of leptin. This paper describes an environmentally friendly and label-free immunosensor based on porous graphene functionalized black phosphorus (PG-BP) composite to detect of leptin. The PG-BP was synthesized via strong coherent coupling between porous graphene (PG) surface plasmons and anisotropic black phosphorus (BP) localized surface plasmons, which made the electrochemical performance of PG and BP synergistic as well as increased the stability and conductive capability of BP material. The PG-BP modified electrodes was further prepared by gold nanoparticles, cysteamine, and glutaraldehyde in turn. Due to the cross-linking effect of glutaraldehyde, anti-leptin can be firmly fixed. These properties of the platform improved the conductive capability of the immunosensor and enhanced the load capacity of the proteins, thereby, the sensitivity of the immunosensor was significantly increased. Under the optimal conditions, the proposed immunosensor exhibited a wide linear range of 0.150-2500 pg/mL with a low detection limit of 0.036 pg/mL. The leptin immunosensor displayed excellent selectivity and anti-interference ability, which could be used for early screening and diagnosis of clinical NALFD.

A novel electrochemical immunosensor based on PG for early screening of depression markers-heat shock protein 70.

In this study, a novel electrochemical immunosensor for early screening of depression markers-heat shock protein 70 (HSP70) was successfully developed based on the porous graphene (PG) with huge specific surface area and excellent structure. Benefiting from the strong adsorption and good bioactivity of PG which was initially prepared via a simple pyrolysis process, a variety of heat shock protein70 (HSP70) can be firmly loaded on the PG to construct the basic electrode (HSP70/PG/GCE)，which was characterized by the cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS), respectively. Due to the HSP70 fixed on the surface of basic electrode and the HSP70 in the samples can competitively combine with the horseradish peroxidase labeled human HSP 70 antibody (HRP-Strept-Biotin-Ab). As a result, it presented a negative correlation between the concentration of HSP70 in samples and the detection signal of the proposed electrochemical immunosensor (HRP-Strept-Biotin-Ab-HSP70/PG/GCE) in the test liquid. The application of PG with excellent electrical conductivity in construction of immunosensor remarkably improved the sensitivity of the immunosensor for detection of HSP70. The proposed immunosensor demonstrated a wide linear range of 0.0448 ~ 100 ng/mL with a low detection limit of 0.02 ng/mL at 3σ. Moreover, the proposed immunosensor could be applied for the sensitive and efficient detection of HSP70 in real samples with good precision, acceptable stability, reproducibility and satisfactory results. Therefore, the HSP70 immunosensor provides a novel and convenient method for early clinical screening of depression markers-heat shock protein 70.

Development of an Analytical Method Based on Temperature Controlled Solid-Liquid Extraction Using an Ionic Liquid as Solid Solvent.

At the present paper, an analytical method based on temperature controlled solid-liquid extraction (TC-SLE) utilizing a synthesized ionic liquid, (N-butylpyridinium hexafluorophosphate, [BPy]PF6), as solid solvent and phenanthroline (PT) as an extractant was developed to determine micro levels of Fe(2+) in tea by PT spectrophotometry. TC-SLE was carried out in two continuous steps: Fe(2+) can be completely extracted by PT-[BPy]PF6 or back-extracted at 80 °C and the two phases were separated automatically by cooling to room temperature. Fe(2+), after back-extraction, needs 2 mol/L HNO3 as stripping agent and the whole process was determined by PT spectrophotometry at room temperature. The extracted species was neutral Fe(PT)mCl2 (m = 1) according to slope analysis in the Fe(2+)-[BPy]PF6-PT TC-SLE system. The calibration curve was Y = 0.20856X - 0.000775 (correlation coefficient = 0.99991). The linear calibration range was 0.10-4.50 µg/mL and the limit of detection for Fe(2+) is 7.0 × 10(-2) µg/mL. In this method, the contents of Fe(2+) in Tieguanyin tea were determined with RSDs (n = 5) 3.05% and recoveries in range of 90.6%-108.6%.