Screening of Acetylcholinesterase Inhibitors by Capillary Electrophoresis with Oriented-Immobilized Enzyme Microreactors Based on Gold Nanoparticles.

A facial and efficient method for the screening of acetylcholinesterase (AChE) inhibitors by capillary electrophoresis was developed. Based on the specific affinity of concanavalin A (Con A) for binding to the glycosyl group of AChE, enzyme molecules were oriented-immobilized on the surface of gold nanoparticles (AuNPs@Con A@AChE). Then, these modified nanoparticles were bounded to the capillary inlet (about 1.0 cm) by electrostatic self-assembly to obtain the oriented-immobilized enzyme microreactor (OIMER). Compared to an IMER with a free enzyme, the peak area of the product obtained by the OIMER increased by 52.6%. The Michaelis-Menten constant (Km) was as low as (0.061 ± 0.003) mmol/L. The method exhibits good repeatability with a relative standard deviation (RSD) of 1.3% for 100 consecutive runs. The system was successfully applied to detect the IC50 values of donepezil and four components from Chinese medicinal plants. This work demonstrates the potential of this method as a low cost, simple, and accurate screening method for other enzyme inhibitors.

Continuous chirped-wave phase-sensitive optical time domain reflectometry.

This Letter proposes a novel phase-sensitive optical time domain reflectometry (Φ-OTDR) with continuous chirped-wave (CCW), which can make full use of both time and frequency domain resources. The principle and benefits of CCW Φ-OTDR are elaborated. With the merit of CCW Φ-OTDR, 1.042 MHz sensing bandwidth and 5pε/Hz strain sensitivity are achieved along a 1013 m fiber with 4.4 m spatial resolution. To the best of the authors' knowledge, this is the first time that a Φ-OTDR achieves megahertz sensing bandwidth with metric spatial resolution, and without limiting the frequency feature of the disturbance. The good performance in long-range sensing is also verified over a 49.7 km fiber. More than that, the digital domain flexibility of the proposed scheme can be used to optimize the measured acoustic signal according to its feature and the practical needs.

Continuous chirped-wave phase-sensitive optical time domain reflectometry: publisher's note.

This publisher's note contains corrections to Opt. Lett.46, 685 (2021)OPLEDP0146-959210.1364/OL.415087.

Quasi-distributed acoustic sensing with interleaved identical chirped pulses for multiplying the measurement slew-rate.

Quasi-distributed acoustic sensing (Q-DAS) based on ultra-weak fiber Bragg grating (UWFBG) is currently attracting great attention, due to its high sensitivity and excellent multiplexing capability. Phase-sensitive optical time-domain reflectometry (Φ-OTDR) based on phase demodulation is one of the most promising interrogation schemes for Q-DAS. In this article, a novel interleaved identical chirped pulse (IICP) approach is proposed on the basis of pulse compression Φ-OTDR with coherent detection. Different from the frequency-division-multiplexing (FDM) method, the identical pulses are used for multiplexing in the IICP scheme, and the mixed reflection signals can be demodulated directly, so the inconsistent phase offsets in FDM can be avoided. As a result, this scheme can enlarge the measurement slew-rate (SR) of Q-DAS by times compared with traditional single pulse scheme. In the proof-of-principle experiment, the SR of 28.9 mɛ/s has been achieved with an 860 m sensing range, which is 5 times as that of the traditional single pulse scheme; meanwhile, the response bandwidth has been enlarged by 5 times. The 277 kHz response bandwidth has been achieved, with 5 m spatial resolution and 2.8 pε/Hz strain sensitivity.

A paper-based colorimetric assay system for sensitive cysteine detection using a fluorescent probe.

A fluorescent probe for the colorimetric detection of cysteine (Cys) was synthesized by combining resorufin with 7-nitrobenzofurazan. The resultant probe was characterized by FT-IR spectroscopy, fluorescence detection, mass spectrometry and NMR spectroscopy. After reacting with Cys, resorufin is released and the color changes from light yellow to red on paper-based analytical devices. The results were recorded using a common cell phone and subsequently analyzed using Photoshop software. The color intensity of the RGB channel increased linearly in the 0.04 to 70.04 µM Cys concentration range when the probe concentration was 2.6 mM (R = 0.9965), and the limit of detection was 16 nM. The effects of detection conditions have been investigated and are discussed. The interference deviations of 13 substances to Cys were in the range of -2% to +2%. The Cys concentration was determined as 270.8 ± 19.3 µM in human plasma, and the recovery was from 99.7% to 100.4%. The work demonstrates the potential of the method to detect Cys in real samples with low cost and high sensitivity.

Colorimetric determination of cysteine by a paper-based assay system using aspartic acid modified gold nanoparticles.

A method is described for cysteine (Cys) determination on paper-based analytical devices using aspartic acid modified gold nanoparticles (Asp-AuNPs). The Asp-AuNPs were characterized by their size, zeta potential, and UV-visible absorption spectrum. After the addition of Cys, it will interact with Asp-AuNPs selectively and leads to the aggregation of Asp-AuNPs. A color change from red to blue can be observed on the paper-based analytical devices. The results were recorded using a cell phone and subsequently analyzed using the Photoshop software. The ratiometric color intensity at red channel and blue channel (Red/Blue) increased linearly in the range 99.9-998.7 µM for Cys (R = 0.9984), and the limit of detection was 1.0 µM. The effects of assay conditions have been investigated and are discussed. The Cys concentration was determined as (0.27 ± 0.02 mM) in human plasma, and the recovery was from 99.2 to 101.1%. Graphical abstract Schematic representation of the paper-based assay system using aspartic acid modified gold nanoparticles (Asp-AuNPs). The ratiometric color intensity method was used for the cysteine (Cys) determination.

Protein C receptor stimulates multiple signaling pathways in breast cancer cells.

The protein C receptor (PROCR) has emerged as a stem cell marker in several normal tissues and has also been implicated in tumor progression. However, the functional role of PROCR and the signaling mechanisms downstream of PROCR remain poorly understood. Here, we dissected the PROCR signaling pathways in breast cancer cells. Combining protein array, knockdown, and overexpression methods, we found that PROCR concomitantly activates multiple pathways. We also noted that PROCR-dependent ERK and PI3k-Akt-mTOR signaling pathways proceed through Src kinase and transactivation of insulin-like growth factor 1 receptor (IGF-1R). These pathway activities led to the accumulation of c-Myc and cyclin D1. On the other hand, PROCR-dependent RhoA-ROCK-p38 signaling relied on coagulation factor II thrombin receptor (F2R). We confirmed these findings in primary cells isolated from triple-negative breast cancer-derived xenografts (PDX) that have high expression of PROCR. To the best our knowledge, this is the first comprehensive study of PROCR signaling in breast cancer cells, and its findings also shed light on the molecular mechanisms of PROCR in stem cells in normal tissue.

Using open-tubular capillary electrochromatography with part-coating column for binding constants determination of ß2 -adrenergic receptor with seven drugs.

An open-tubular capillary electrochromatography method has been developed for the determination of binding constants between ß2 -adrenergic receptor (ß2 -AR) and seven drugs. ß2 -AR was oriented immobilized onto one part of inner surface of capillary via microwave-assisted technical synthesis. According to the linear relationship between coating length and the apparent mobility of analyte, the binding constant (Kb ) can be obtained by related theories and equations. The order of Kb values between drugs such as adrenaline hydrochloride, norepinephrine bitartrate, and propranolol hydrochloride with ß2 -AR is well consistent with that reported in the literature. By the method, Kb values between four extracts of Radix Paeoniae Rubra and ß2 -AR were also successfully obtained. Subsequently, computer models were applied to interpret the CEC experiments. And the results proved to be in good agreement with the method. The work, herein, demonstrates the potential of the method in drug-receptor affinity interactions evaluation and screening of lead compounds from natural sources.

Enantiomeric separation of adrenaline, noradrenaline, and isoprenaline by capillary electrophoresis using streptomycin-modified gold nanoparticles.

Enantiomeric separations of the adrenergic compounds adrenaline, noradrenaline, and isoprenaline were studied. Electromigrative separations were performed in uncoated fused silica capillaries using streptomycin-modified gold nanoparticles (ST-AuNPs) as an additive to the background electrolyte. The ST-AuNPs are shown to serve as an effective chiral selector. The modified AuNPs were characterized in terms of size and zeta potential, and by IR and UV-vis spectra. The effects of ST-AuNP concentration, pH value, temperature, and separation voltage on the separations were systematically studied. Under optimized experimental conditions, racemic mixtures of the respective adrenergic drugs were baseline-separated within 7 min with a resolution of up to 7.5. The relative standard deviations of the resolution in inter-day and intra-day studies (n = 5) were generally <5%. Graphical abstract Schematic of the method for enantiomeric separations. (A): At low concentrations of streptavidinylated gold nanoparticles (ST-AuNPs), the better matching enantiomer is preferably "transported" by the ST-AuNPs; (B) ST-AuNP concentration increased to an optimal value; (C): The ST-AuNP concentration is too high; even poorly matching enantiomers will be transported simultaneously.

A microfluidic paper-based device to assess acetylcholinesterase activity.

Neurotransmitters play key roles in cell-to-cell communication. These chemical messengers are involved in many functional processes, including growth, reproduction, memory, and behavior. In this communication, we describe a novel microfluidic paper-based analytical device (µPAD) to detect acetylcholinesterase (AChE) activity and inhibitor screening through a colorimetric analysis. The µPAD is easily fabricated via a wax printing process whereby wax is deposited onto the surface of chromatographic paper, and heated to create a hydrophobic barrier. Separate solutions of 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) and samples containing AChE and acetylthiocholine iodide (ATC) (or cysteine, Cys), respectively, are directly spotted onto the µPAD. DTNB and AChE/ATC (or Cys) flow towards each other where a reaction occurs to form the yellow colored 2-nitro-5-thiobenzoic acid anion (TNB2- ). The device is dried, scanned, and analyzed yielding a linear range of average inverse yellow intensities versus substrate concentration. An IC50 value (0.045 nM) with a known inhibitor, neostigmine bromide (NB), is obtained on the device. µPADs are low cost and easy to fabricate and have great potential to quantify neurotransmitter activity.

A Colorimetric Sensor for the Highly Selective Detection of Sulfide and 1,4-Dithiothreitol Based on the In Situ Formation of Silver Nanoparticles Using Dopamine.

Hydrogen sulfide (H2S) has attracted attention in biochemical research because it plays an important role in biosystems and has emerged as the third endogenous gaseous signaling compound along with nitric oxide (NO) and carbon monoxide (CO). Since H2S is a kind of gaseous molecule, conventional approaches for H2S detection are mostly based on the detection of sulfide (S2-) for indirectly reflecting H2S levels. Hence, there is a need for an accurate and reliable assay capable of determining sulfide in physiological systems. We report here a colorimetric, economic, and green method for sulfide anion detection using in situ formation of silver nanoparticles (AgNPs) using dopamine as a reducing and protecting agent. The changes in the AgNPs absorption response depend linearly on the concentration of Na2S in the range from 2 to 15 µM, with a detection limit of 0.03 µM. Meanwhile, the morphological changes in AgNPs in the presence of S2- and thiol compounds were characterized by transmission electron microscopy (TEM). The as-synthetized AgNPs demonstrate high selectivity, free from interference, especially by other thiol compounds such as cysteine and glutathione. Furthermore, the colorimetric sensor developed was applied to the analysis of sulfide in fetal bovine serum and spiked serum samples with good recovery.

Research on drug-receptor interactions and prediction of drug activity via oriented immobilized receptor capillary electrophoresis.

Oriented covalent immobilized ß2 -adrenergic receptor (ß2 -AR) CE (OIRCE) was developed to determine the interactions between a set of natural extracts of Radix Paeoniae Rubra (NERPR) and ß2 -AR, and to predict the activity of NERPR. The inner capillary surface is chemically bonded with stable ß2 -AR coating via microwave-assisted technical synthesis. The modified capillaries were characterized via infrared spectroscopy and fluorescence microscopy. Furthermore, the bonding amounts of ß2 -AR were first obtained via fluorescence spectroscopy method. In determining the amount of bonded ß2 -AR, the regression equation A  =  576 707C + 35.449 and the correlation coefficient 0.9995 were obtained. This result revealed an excellent linear relationship in the range of 2 × 10(-4)  mg/mL to 1 × 10(-3)  mg/mL. The normalized capacity factor (KRCE ) was obtained using OIRCE in evaluating drug-receptor interactions. Related theories and equations were used to calculate KRCE values from apparent migration times of a solute and EOF. The order of KRCE and the binding constant (Kb ) values between drugs and ß2 -AR was well consistent. The results confirmed that the OIRCE and KRCE values can be effectually used to investigate drug-receptor interactions, and OIRCE has the potential to predict drug activity and to select leading compounds from natural chemicals.

[Purification Technology Optimization for Saponins from Ziziphi Spinosae Semen with Macroporous Adsorption Resin by Box-Behnken Design-Response Surface Methodology].

OBJECTIVE: To compare the purification effect of saponins from Ziziphi Spinosae Semen with different types of macroporous adsorption resin, and to optimize its purification technology. METHODS: The type of macroporous resins was optimized by static adsorption method. The optimum technological conditions of saponins from Ziziphi Spinosae Semen was screened by single factor test and Box-Behnken Design-Response Surface Methodology. RESULTS: AB-8 macroporous resin had better purification effect of total saponins than other resins, optimum technological parameters were as follows: column height-diameter ratio was 5: 1, the concentration of sample solution was 2. 52 mg/mL, resin adsorption quantity was 8. 915 mg/g, eluted by 3 BV water, flow rate of adsorption and elution was 2 BV/h, elution solvent was 75% ethanol, elution solvent volume was 5 BV. CONCLUSION: AB-8 macroporous resin has a good purification effect on jujuboside A. The optimized technology is stable and feasible.

Improving pesticide residue detection: Immobilized enzyme microreactor embedded in microfluidic paper-based analytical devices.

Orientationally immobilized enzyme microreactors (OIMERs), embedded in microfluidic paper-based analytical devices (µPADs) were developed for improved detection of pesticide residues in food. Acetylcholinesterase (AChE) was orientationally immobilized on the reusable Part I of the µPADs, using the specific affinity binding of concanavalin A (Con A) to a glycosyl group on AChE. Using the disposable Part II, facile colorimetric quantification was performed with a smartphone and software, or qualitative detection by a naked-eye visual test. The AChE immobilized in OIMERs not only had improved activity and stability, but also high sensitivity, with a limit of detection as low as (0.007 ± 0.003) µg/mL. The method was used to detect pesticides residues in real vegetable samples; the recovery (88.6-102.7%) showed high reliability for pesticide residues detection in foods. A molecular docking study and an enzyme kinetic analysis were conducted to characterize the mechanism of action of the OIMERs.

Facial on-line enrichment of glycoproteins by capillary electrophoresis with boronate-functionalized poly(glycidyl methacrylate) microparticles coated column.

A facial and rapid method for glycoproteins enrichment by capillary electrophoresis was developed. The 3-aminophenylboronic acid-functionalized poly(glycidyl methacrylate) microparticles (PGMA@APBA) were attached to the capillary inlet (length of ∼1.5 cm) by electrostatic self-assemble action to prepare a partially coated capillary column. The process is simple and reversible, allowing for easy renewal of the PGMA@APBA coating when its enrichment efficiency decreases. By utilizing the coated column, glycoproteins can be enriched within 2 min. The column exhibits a specific enrichment for glycoproteins and can be consecutively used for approximately 60 runs. The relative standard deviations (RSDs) of peak area of run-to-run (n = 5) and batch-to-batch (n = 3) were 1.5 % and 1.0%, respectively. The method was successfully applied to enrich glycoproteins from 1 × 1012-fold diluted real egg white sample, indicating its practical applicability.

Niche inflammatory signals control oscillating mammary regeneration and protect stem cells from cytotoxic stress.

Stem cells are known for their resilience and enhanced activity post-stress. The mammary gland undergoes frequent remodeling and is subjected to recurring stress during the estrus cycle, but it remains unclear how mammary stem cells (MaSCs) respond to the stress and contribute to regeneration. We discovered that cytotoxic stress-induced activation of CD11c+ ductal macrophages aids stem cell survival and prevents differentiation. These macrophages boost Procr+ MaSC activity through IL1ß-IL1R1-NF-κB signaling during the estrus cycle in an oscillating manner. Deleting IL1R1 in MaSCs results in stem cell loss and skewed luminal differentiation. Moreover, under cytotoxic stress from the chemotherapy agent paclitaxel, ductal macrophages secrete higher IL1ß levels, promoting MaSC survival and preventing differentiation. Inhibiting IL1R1 sensitizes MaSCs to paclitaxel. Our findings reveal a recurring inflammatory process that regulates regeneration, providing insights into stress-induced inflammation and its impact on stem cell survival, potentially affecting cancer therapy efficacy.

Drag Reduction Technology of Water Flow on Microstructured Surfaces: A Novel Perspective from Vortex Distributions and Densities.

Revealing the turbulent drag reduction mechanism of water flow on microstructured surfaces is beneficial to controlling and using this technology to reduce turbulence losses and save energy during water transportation. Two microstructured samples, including a superhydrophobic and a riblet surface, were fabricated near which the water flow velocity, and the Reynolds shear stress and vortex distribution were investigated using a particle image velocimetry. The dimensionless velocity was introduced to simplify the Ω vortex method. The definition of vortex density in water flow was proposed to quantify the distribution of different strength vortices. Results showed that the velocity of the superhydrophobic surface (SHS) was higher compared with the riblet surface (RS), while the Reynolds shear stress was small. The vortices on microstructured surfaces were weakened within 0.2 times that of water depth when identified by the improved ΩM method. Meanwhile, the vortex density of weak vortices on microstructured surfaces increased, while the vortex density of strong vortices decreased, proving that the reduction mechanism of turbulence resistance on microstructured surfaces was to suppress the development of vortices. When the Reynolds number ranged from 85,900 to 137,440, the drag reduction impact of the superhydrophobic surface was the best, and the drag reduction rate was 9.48%. The reduction mechanism of turbulence resistance on microstructured surfaces was revealed from a novel perspective of vortex distributions and densities. Research on the structure of water flow near the microstructured surface can promote the drag reduction application in the water field.

The Roles of Riblet and Superhydrophobic Surfaces in Energy Saving Using a Spatial Correlation Analysis.

Riblet and superhydrophobic surfaces are two typical passive control technologies used to save energy. In this study, three microstructured samples-a micro-riblet surface (RS), a superhydrophobic surface (SHS), and a novel composite surface of micro-riblets with superhydrophobicity (RSHS)-were designed to improve the drag reduction rate of water flows. Aspects of the flow fields of microstructured samples, including the average velocity, turbulence intensity, and coherent structures of water flows, were investigated via particle image velocimetry (PIV) technology. A two-point spatial correlation analysis was used to explore the influence of the microstructured surfaces on coherent structures of water flows. Our results showed that the velocity on microstructured surface samples was higher than that on the smooth surface (SS) samples, and the turbulence intensity of water on the microstructured surface samples decreased compared with that on the SS samples. The coherent structures of the water flow on microstructured samples were restricted by length and structural angles. The drag reduction rates of the SHS, RS, and RSHS samples were -8.37 %, -9.67 %, and -17.39 %, respectively. The novel established RSHS demonstrated a superior drag reduction effect and could improve the drag reduction rate of water flows.

LRP12 is an endogenous transmembrane inactivator of α4 integrins.

Dynamic regulation of integrin activation and inactivation is critical for precisely controlled cell adhesion and migration in physiological and pathological processes. The molecular basis for integrin activation has been intensively studied; however, the understanding of integrin inactivation is still limited. Here, we identify LRP12 as an endogenous transmembrane inhibitor for α4 integrin activation. The LRP12 cytoplasmic domain directly binds to the integrin α4 cytoplasmic tail and inhibits talin binding to the ß subunit, thus keeping integrin inactive. In migrating cells, LRP12-α4 interaction induces nascent adhesion (NA) turnover at the leading-edge protrusion. Knockdown of LRP12 leads to increased NAs and enhanced cell migration. Consistently, LRP12-deficient T cells show an enhanced homing capability in mice and lead to aggravated chronic colitis in a T cell-transfer colitis model. Altogether, LRP12 is a transmembrane inactivator for integrins that inhibits α4 integrin activation and controls cell migration by maintaining balanced NA dynamics.

Interactions between isoprenaline hydrochloride and bovine serum albumin (BSA) in capillary zone electrophoresis and affinity capillary electrophoresis.

Capillary zone electrophoresis (CZE) and affinity capillary electrophoresis (ACE) were developed to investigate the interactions between isoprenaline hydrochloride (IH) and bovine serum albumin (BSA). In CZE, the binding constant (Kb) was 4.07 x 10(4) M(-1) (298 K) and 4.76 x 10(4) M(-1) (310K) using the Scatchard analysis. The number of binding sites (n) in this interaction was approximately one (n approximately equal 1). Furthermore, thermodynamic parameters, such as changes in Gibbs free energy (deltaG), enthalpy (deltaH), and entropy (deltaS) of the binding procedure were also obtained. At 298 K, deltaG, deltaH, and deltaS were -26.30 kJ x mol(-1), 10.02 kJ x mol(-1), and 0.12 kJ x mol(-) x K(-1), respectively. The deltaG at 310K was -27.76 kJ x mol(-1), whereas the deltaH and deltaS at 310 K were identical with that at 298 K. In ACE, a more reliable parameter, mobility ratio (M), was applied in the calculation of Kb. Kb (310K) obtained through this method was 9.80 x 10(4) and 9.24 x 10(4) M(-1) when IH and BSA were added to the buffer in varying concentrations, respectively. The obtained Kb may help in gaining some insights on the possible drug/protein interactions and in the early evaluation of the pharmacokinetic profile of the drug during cardiovascular drug screening.