Determination of Microamounts of Proteins by Resonance Light Scattering with Copper Phthalocyanine Tetrasulfonic Acid

Abstract

Based on the strong enhancement effect of proteins on the resonance light scattering of copper phthalocyanine tetrasulfonic acid, a method for the determination of microamounts of proteins has been developed. Under the experimental conditions (2.0×10^?6 mol/L copper phthalocyanine tetrasulfonic acid, pH 2.60, ionic strength 0.001 mol/L NaCl), the linear range of this assay is 0.06–4.0 µg/mL for bovine serum albumin (BSA), 0.1–2.0 µg/mL for human serum albumin (HSA), 0.0–2.0 µg/mL for human γ‐IgG, and 0.2–6.0 µg/mL for ovalbumin. The detection limits (3δ) are 16.8 ng/mL for BSA, 23.4 ng/mL for HSA, 37.6 ng/mL for human γ‐IgG, and 48.3 ng/mL for ovalbumin, respectively. This method has been applied to the analysis of total proteins in human serum samples collected from the hospital, and the results were in good agreement with those reported by the hospital.     

Gold nanoparticle-based signal augmentation of quartz crystal microbalance immunosensor measuring C-reactive protein

A biotinylated anti-rat C-reactive protein (CRP) antibody was ingeniously prepared by the reaction of the unmodified antibody with a water-soluble sulfosuccinimidyl-6-(biotinamido)hexanoate. The molar biotin incorporation of the resulting modified antibody was found as 5.82. A flow-type indirect-competitive quartz crystal microbalance immunosensor system was constructed with the sensor chip immobilized with 2 mg/mL CRP as the coating antigen. When 200 μL of the modified antibody having the concentration of 0.250 mg/mL was added with a streptavidin-coated gold nanoparticle (GNP) to the immunosensor system, the frequency shift obtained was 139.8 ± 0.3 Hz. Compared to the frequency shift of 91.1 ± 1.3 Hz found with the addition of the unmodified antibody only, the signal augmentation after GNP binding amounted to 53.4%, which resulted in sensitivity improvement of the current immunosensor.     

E-glass fibers coated with nickel phosphorous by electroless deposition technique

The nickel phosphorous catalyst coating was studied on the E-glass fiber by electroless deposition using sodium hypophosphite as a reducing agent in alkaline medium. The effects of deposition time, deposition temperature, pH of the solution, and amount of stabilizer used in the solution on the coating efficiency were investigated by X-ray diffraction (XRD), energy dispersive X-ray analysis (EDAX), scanning electron microscopy (SEM), thermogravimetric analysis (TGA), and differential thermal analysis (DTA). The XRD indicates the presence of amorphous and crystalline phases of Ni and nickel–phosphorous (NiP). From TGA, it is evident that the NiP coating increases the thermal stability and magnetic properties of the glass fiber. The bath temperature of 80 °C, pH of 9 to 9.5, and stabilizer concentration of 25 g/l are optimum to get a good and uniform coating of NiP on glass fiber. Carbon nanotubes were successfully coated on NiP coated glass fiber at a temperature of 700 °C.     

Application of mesoporous carbon and modified mesoporous carbon for treatment of DMF sewage

Mesoporous carbon (MC) was prepared in soft template, and potassium ferricyanide was added into MC to prepare the modified mesoporous carbon (MMC). TEM, SEM, FT-IR, and N₂ adsorption–desorption were used to characterize the textural properties of mesoporous materials. The BET specific surface area, pore volume, and the pore size of MC and MMC were 607.6321 and 304.7475 m²/g, 0.313552 and 0.603573 cm³/g, and 5.4356 and 7.9227 nm, respectively. The adsorption capabilities of MC and MMC were compared with the silica mesoporous material MCM-41. The influences of different adsorption conditions were optimized. For MC, the optimums of adsorbent dose, DMF initial concentration, rotating speed, and pH were 0.002 mg/50 mL, 200 mg/L, 200 r/min, and 4, respectively. MMC showed the highest DMF adsorption capacity at adsorbent dose 0.002 g/50 mL, DMF initial concentration 1000 mg/L, rotating speed 1000 r/min, pH more than 9, and contact time of less than 20 min. Meanwhile for MC, MMC, Pseudo-second-order equation was used to fit adsorption kinetics data. And adsorption process could be well fitted by Langmuir and Freundlich adsorption isotherms of MC, MMC. The results showed that MMC was a perfect adsorbent for DMF, and it was easy to separation and recycle. The recycling property of MMC was still relatively better than other two adsorbents.     

Development of Dissociation-Enhanced Lanthanide Fluoroimmunoassay for Measuring Leptin

Development of a dissociation-enhanced lanthanide fluoroimmunoassay (DELFIA) for measuring leptin, a satiety hormone of appetite control, was conducted in sandwich assay format exploiting a microplate immobilized with an anti-leptin antibody and another antibody raised against leptin and tagged with an europium chelate. In the leptin DELFIA of this study, amounts of antibody coated to the microplate and of the bioconjugate for the second immune reaction were optimized as 0.5 μg and 200 ng per well, respectively. When plotted in double-logarithmic scale, a linear relationship of y (log₁₀ response signal) = 0.6023× (log₁₀ leptin concentration) + 3.4084 (r² = 0.9646) was obtained at the leptin concentrations of 0.01─50 ng/mL with the limit of detection of 0.01 ng/mL. Individual leptin concentrations in various samples were well convergent to the calibration curve of the current assay. When applied to the measurement of leptin in a rat serum, the present assay was found quite effective and was competitive to a commercial sandwich-type ELISA.     

Lipid-based nanoparticles as drug delivery system for paclitaxel in breast cancer treatment

Paclitaxel (PTX) is a well-known antitumor drug, widely utilized in the treatment of breast, ovarian, head, and neck tumors, among others. The low aqueous solubility (< 1.0 μg/mL; log P = 3.96) limits its use by intravenous route, and alternatives found for the marketed products are associated with high toxicity. Incorporation of PTX into lipid nanocarriers has been considered an interesting nontoxic alternative for this route, but drug loading is usually low. This study aimed to analyze the influence of the lipid composition and three different lipid nanosystems—solid lipid nanoparticles, nanostructured lipid carriers (NLCs), and nanoemulsion—in PTX encapsulation and its biological response. The three proposed systems were prepared by hot melt homogenization followed by ultrasonication. Among the blank formulations first prepared, NLC had the smallest size (74 ± 1 nm), with negative zeta potential (? 11.4 ± 0.1 mV). The incorporation of 0.10 mg/mL PTX into this NLC formulation yielded high and stable encapsulation (0.089 ± 0.003 mg/mL), also supported by polarized light microscopy and differential scanning calorimetry curves. NLC-PTX was very effective against MCF-7 (IC50 25.33 ± 3.17 nM) and MDA-MB-231 breast cancer cell lines (IC50 2.13 ± 0.21 nM), compared to free PTX (IC50 > 500 nM). In addition, no significant cytotoxicity was found against fibroblast cells. Taken together, these results demonstrated that PTX was successfully incorporated into NLC with appropriate physicochemical characteristics for intravenous administration, suggesting that the use of NLC as vehicle to incorporate PTX may be a promising strategy in the treatment of breast cancer.     

Facile preparation of magnetically functionalized graphite nanosheets for porcine pancreatic lipase immobilization

We report that silicon nanoparticles (SiNPs) with typical sizes from 5 to 50 nm prepared by grinding of porous silicon can act as efficient scavengers of human immunodeficiency virus (HIV) and respiratory syncytial virus (RSV). In vitro studies have revealed a strong suppression of the viral activity in the presence of SiNPs with concentration above 0.1 and 0.01 mg/mL for HIV and RSV, respectively. The observed effect is explained by binding of the virions with SiNPs that is supposed to be universal for different enveloped viruses. Because of the cytotoxic concentration of SiNPs is of the order of 1 mg/mL, SiNPs can be proposed for applications in new harmless methods of antiviral treatment.     

Liquid Chromatography Coupled with Fluorimetric Detection and Third Derivative Synchronous Fluorescence Spectroscopy as two Analytical Methods for the Simultaneous Determination of Rabeprazole Sodium and Domperidone After Derivatization with 4-Chloro-7-Nitrobenzofurazan

Two simple, sensitive, rapid, economic and validated methods, namely reversed phase liquid chromatography (method Ι) and third derivative synchronous fluorescence spectroscopy (method ΙΙ) have been developed for the simultaneous determination of rabeprazole sodium and domperidone in their laboratory prepared mixture after derivatization with 4-Chloro-7-nitrobenzofurazan. Reversed phase chromatography was conducted using a Zorbax® SB-Phenyl column (250.0 mm × 4.6 mm id) combined with a guard column at ambient temperature with fluorimetric detection at 540 nm after excitation at 483 nm. A mobile phase composed of a mixture of distilled water with methanol and acetonitrile in a ratio of 50:20:30 adjusted pH to 4 has been used at a flow rate of 1 mL/min. Sharp well resolved peaks were obtained for domperidone and rabeprazole sodium with retention times of 5.5 and 6.4 min respectively. While in method ΙΙ, the third-derivative spectra were estimated at 507 and 436 nm for rabeprazole sodium and domperidone respectively. Linearity ranges for rabeprazole sodium and domperidone respectively in both methods were found to be 0.15–2.0 and 0.1–1.5 μg/mL. The proposed methods were successfully applied for the analysis of the two compounds in their binary mixtures, and laboratory prepared tablets. The obtained results were favorably compared with those obtained by the comparison method. Furthermore, detailed validation procedure was also conducted.     

The effect of high hydrostatic pressure on the survival of Saccharomyces cerevisiae in model suspensions and beetroot juice

The inactivation of Saccharomyces cerevisiae NCFB 3191 using high hydrostatic pressure of 300 MPa at 20°C with a holding time of 0, 1, 5 and 10 min was investigated with model suspensions in phosphate-buffered saline and in beetroot juice. The reduction in S. cerevisiae NCFB 3191 in model suspensions was about 5 log after 10 min of pressurization, irrespective of the initial level of cell concentration in the samples (5.4–8.7 log cfu/mL). The baroprotective effect of beetroot juice on yeast cells during pressurization was observed; the reduction was lower and was only 3.5 log (the inoculum was 5.4 log cfu/mL). No sublethal injury among the surviving cells of the studied yeast strain was found.     

Glass silanization with blocked-isocyanate for the fabrication of DNA microarrays

Microarray glass slides were functionalized with isocyanate-γ-propyltrimethoxysilane and characterized by atomic force microscopy, contact angle measurements. The coating was also tested in microarray DNA hybridization experiments using fluorescence microscopy for the detection of the spots. Isocyanate groups were blocked with both methyl ethyl ketoxime (MEKO) or bisulfite (BS). Contact angle measurements showed that BS blocking agent makes glass slide more hydrophilic than MEKO one likely because of the presence of ionic groups. Atomic force microscopy shows the presence of a continuous domain with some depressed areas that could be attributed to silanized and unmodified glass, respectively. Binding tests for amino-functionalized oligonucleotides did not evidence any difference between the two blocking systems. In contrast to these results, the hybridization efficiency of the two systems are different, since probes spotted on MEKO blocked slides show a fluorescence intensity which is about three times higher than those achieved with BS slides.MEKO blocked glass surface gave very interesting results in terms of ease of preparation and use, sensitivity and selectivity in binding and hybridization as well as a low background noise.     

Preparation of graphene-glass fiber-resin composites and its electromagnetic shielding performance

The surface of the glass fiber (GF) was modified by silane coupling agent (KH550) and bovine serum albumin (BSA), and then the graphene oxide (GO) was coated onto the modified surface of the glass fiber. Followed by a reduction reaction, the reduced graphene oxide (RGO) coated on glass fiber was obtained. Finally, the reduced graphene oxide-glass fibers (RGO-GF) were combined with unsaturated resins. The interfacial morphology of reduced graphene oxide-glass fibers was investigated by scanning electron microscopy (SEM). The structure of the materials was analyzed by Fourier transform infrared spectroscopy (FT-IR). The crystal phases of the material were identified by X - ray diffraction (XRD). The mechanical properties and electromagnetic shielding effectiveness of the sample were tested. The results showed that the interface between glass fibers and graphene binds more closely after the glass fibers was treated by KH550. The tensile strength of the RGO-GF composites reached 85.05 MPa. Compared with the GF composites, it increased by 51.4% when the glass fibers content was 30%. The shielding effectiveness of the composites reached 21.3 dB at the frequency range of 8.2–12.4 GHz (x-band). Therefore, by coating the surface with reduced graphene oxide, the glass fibers can make a great shielding effect on the electromagnetic wave.     

Binding Study of Phenformin with Bovine Serum Album Using a Combined Technique of Equilibrium Dialysis with Flow-Injection Chemiluminescence Detection

ABSTRACT

The interaction between phenformin hydrochloride and bovine serum albumin (BSA) was investigated by the methods of chemiluminescence combined with equilibrium dialysis technique. A novel N-bromosuccinimide (NBS)–eosin Y (EY) chemiluminescence (CL) method was established for the determination of phenformin. The mechanism of this chemiluminescence system was proposed. Optimization studies were performed to determine the phenformin. Under the optimal conditions, the CL intensity was linear for a phenformin concentration over the range of 4.6 × 10^?8 to 5.0 × 10^?5 g/mL. The detection limit was 1.5 × 10^?8 g/mL. The data obtained by the present equilibrium dialysis–CL system were analyzed using the Klotz plot and the Scatchard analysis. The results showed that the Klotz plot and the Scatchard plot are linear with good correlation coefficient, indicating that the phenformin has only one type of binding site on BSA. The binding parameters were the number of the binding sites n (1.02) and the estimated association constant K (2.66 × 10⁴ L/mol). The chemiluminescence system combined with equilibrium dialysis developed in this work demonstrated its use for determination of interaction between drug and protein by using relatively simple instrument.     

MRI contrast agent for molecular imaging of the HER2/neu receptor using targeted magnetic nanoparticles

In this study, Trastuzumab modified Magnetic Nanoparticles (TMNs) were prepared as a new contrast agent for detecting HER2 (Human epidermal growth factor receptor-2) expression tumors by magnetic resonance imaging (MRI). TMNs were prepared based on iron oxide nanoparticles core and Trastuzumab modified dextran coating. The TMNs core and hydrodynamic size were determined by transmission electron microscopy and dynamic light scattering. TMNs stability and cytotoxicity were investigated. The ability of TMNs for HER2 detection were evaluated in breast carcinoma cell lines (SKBr3 and MCF7 cells) and tumor-bearing mice by MRI and iron uptake determination. The particles core and hydrodynamic size were 9 ± 2.5 and 41 ± 15 nm (size range: 15?C87 nm), respectively. The molar antibody/nanoparticle ratio was 3.1?C3.5. TMNs were non-toxic to the cells below the 30 ??g (Fe)/mL concentration and good stable up to 8 weeks in PBS buffer. TMNs could detect HER2 oncogenes in the cells surface with imagable contrast by MRI. The invivo study in mice bearing tumors indicated that TMNs possessed a good diagnostic ability as HER2 specific contrast agent by MRI. TMNs were demonstrated to be able to selectively accumulate in the tumor cells, with a proper signal enhancement in MRI T2 images. So, the complex may be considered for further investigations as an MRI contrast agent for detection of HER2 expression tumors in human.     

Simultaneous determination of amlodipine besylate and rosuvastatin calcium in binary mixtures by voltammetric and chromatographic techniques

Voltammetric and liquid chromatographic (LC) methods have been developed for the simultaneous determination of amlodipine besylate (AML) and rosuvastatin calcium (ROS) for the first time. Detailed electrochemical behavior and simultaneous voltammetric determination of AML and ROS were investigated in detail using glassy carbon electrode (GCE). High-performance liquid chromatography (HPLC) and ultra-performance liquid chromatography (UPLC) were also developed for the comparison. Voltammetric method exhibited linear dynamic responses for the simultaneous assay of AML and ROS in the concentration range between 0.006 and 2.85 μg/mL and between 0.01 and 5.00 μg/mL, with detection limits of 0.001 and 0.003 μg/mL, respectively. On the other hand, LC methods presented a wider linearity range than that of the SWV method between 0.5 and 100 μg/mL with the detection limits of 0.011 and 0.027 μg/mL for AML and 0.034 and 0.042 μg/mL for ROS by UPLC and HPLC techniques, respectively.     

A Novel Homogeneous Time-Resolved Fluoroimmunoassay for Carcinoembryonic Antigen Based on Water-Soluble Quantum Dots

Quantum dots are not widely used in clinical diagnosis. However, the homogeneous time-resolved fluorescence assay possesses many advantages over current methods for the detection of carcinoembryonic antigen (CEA), a primary marker for many cancers and diseases. Therefore, a novel luminescent terbium chelates- (LTCs) and quantum dots-based homogeneous time-resolved fluorescence assay was developed to detect CEA. Glutathione-capped quantum dots (QDs) were prepared from oil-soluble QDs with a 565 nm emission peak. Conjugates (QDs-6 F11) were prepared with QDs and anti-CEA monoclonal antibody. LTCs were prepared and conjugates (LTCs-S001) were prepared with another anti-CEA monoclonal antibody. The fluorescence lifetime of QDs was optimized for sequential analysis. The Förster distance (R₀) was calculated as 61.9 Å based on the overlap of the spectra of QDs-6 F11 and LTCs-S001. Using a double-antibody sandwich approach, the above antibody conjugates were used as energy acceptor and donor, respectively. The signals from QDs were collected in time-resolved mode and analyzed for the detection of CEA. The results show that the QDs were suitable for time-resolved fluoroassays. The spatial distance of the donor-acceptor pair was calculated to be 61.9 Å. The signals from QDs were proportional to CEA concentration. The standard curve was LogY?=?2.75566?+?0.94457 LogX (R?=?0.998) using the fluorescence counts (Y) of QDs and the concentrations of CEA (X). The calculated sensitivity was 0.4 ng/mL. The results indicate that water-soluble QDs are suitable for the homogenous immunoassay. This work has expanded future applications of QDs in homogeneous clinical bioassays. Furthermore, a QDs-based homogeneous multiplex immunoassay will be investigated as a biomarker for infectious diseases in future research.     

Study of an anisotropic ferrimagnetic bioactive glass ceramic for cancer treatment

For the hyperthermia therapy of cancer, ferrimagnetic glass ceramics are a potential candidate. Ferrimagnetic zinc-ferrite-containing bioactive glass ceramics were prepared by quenching the glass ceramics from sintering temperature. Then the samples were heated to 600°C and cooled in an aligning magnetic field of 1 Tesla to cause anisotropy. The magnetically aligned samples were compared with non-aligned samples. Vibrating sample magnetometry measurements at 10 kOe showed that the magnetic properties were enhanced by the aligning magnetic field and it led to an enhancement of the magnetic heat generation under a magnetic induction furnace operating at 500 Oe and 400 kHz for 2 min. Data showed that the maximum specific power loss and temperature increase after 2 min were 31.5 W/g and 45°C, respectively, for the aligned sample of maximum zinc-ferrite crystalline content. The glass ceramics were immersed in simulated body fluid for 3 weeks. X-ray diffraction and Fourier transform infrared and atomic absorption spectroscopy results indicated the growth of precipitated hydroxyapatite, suggesting that the ferrimagnetic glass ceramics were bioactive and could bond to living tissues in physiological environment.     

Thermally Stable Schiff Base and its Metal Complexes: Molecular Docking and Protein Binding Studies

In this paper, interaction of Schiff base and its metal complexes carrying naphthalene ring in the structure with bovine serum albumin (BSA) were investigated using UV-vis absorption, fluorescence spectroscopies and molecular docking methods. The effect on the binding mechanism and properties of these compounds containing metal-free, iron and copper ions were also investigated. The fluorescence spectroscopy results showed that fluorescence intensity of BSA in the presence of different concentration of ligands was decreased through a static quenching mechanism. Binding constants (KSV, Kbin and Ka) and thermodynamic parameters (ΔG, ΔH and ΔS) for the ligand-protein interactions were also determined. ΔG values of ligand-protein interaction were calculated in the range ? 6.3 to ?5.5 kcal/mol. These negative values showed that binding process is spontaneous and, hydrogen bonding and van der Waals force were main interaction of the protein and ligands. ΔH and ΔS value were also calculated in the range of 1.10 to 1.26 kJ/mol and 0.133 to 0.135 kJ/mol. K, respectively. These positive values indicated that the binding process between ligands and BSA are endothermic and electrostatic interaction, respectively.     

Detection of the potential tumor marker of AFP using surface‐enhanced Raman scattering‐based immunoassay

The development of rapid, highly sensitive detection methods for α‐fetoprotein (AFP) is very important. As hepatocellular carcinoma is closely related to the level of AFP in the blood, it is necessary to maintain an AFP concentration below the safety limit. In this paper, we propose a universal, rapid, sensitive, and highly specific immunoassay system utilizing gold nanoparticles (AuNPs) and surface‐enhanced Raman scattering (SERS). This new system features a sandwich structure combining mercaptobenzoic acid‐labeled immunogold nanoparticles with the antigen and the antibody atop a pre‐designed substrate made of a glass slide modified with AuNPs. This SERS‐based immunoassay can detect AFP concentrations as low as 100 pg/ml, which is a significant improvement on the capabilities of the enzyme‐linked immunosorbent assay method. A good linear relationship between the SERS peak intensity and the logarithm of antigen concentrations (from 1 ng/ml to 100 ng/ml) was observed. This technique provides an effective model for the detection of biomarkers in medical diagnostics, criminal investigation, and other fields. Copyright © 2013 John Wiley & Sons, Ltd.     

Novel Fluorescent Membrane for Metronidazole Sensing Prepared by Covalent Immobilization of a Pyrenebutyric Acid Derivative

In the present paper, we report the fabrication of a new sensing membrane for fluorescence detection of metronidazole (MNZ). Briefly, a pyrenebutyric acid derivative, 2-(methacryloyloxy) ethyl-4-(1-pyrenyl) butanoate (MPB) with a double bond, was synthesized and copolymerized with 2-hydroxyethylmethacrylate (HEMA) on the activated glass surface by thermal initiation in the presence of cross-linker. The sensor responds linearly to metronidazole in the concentration range of 1.23~35.48 mg.L^?1 in aqueous solution with a detection limit of 0.36 mg.L^?1. The lifetime is enhanced by covalently immobilizing the pyrenebutyric acid derivative on glass slide, which hinders leaching of the dye from the membrane. The sensor could be regenerated after use by washing in methanol (RSD?=?2.42 %), and it shows sufficient stability, and selectivity. Interference of other pharmaceuticals on membrane performance is discussed. The developed membrane has been successfully applied for the direct determination of metronidazole in human serum sample without pretreatment.     

A strategy for immunoassay signal amplification using clusters of immunogold nanoparticles

The clusters of immunogold nanoparticles were fabricated through cross-linking immunogold nanoparticles with glutaraldehyde. A novel strategy for immunoassay signal amplification using the clusters of immunogold nanoparticles on glass slides based on electroless deposition was described. The immunoassay signal amplification through the clusters of immunogold nanoparticles was about three to four times higher than that through single immunogold nanoparticles. The purple attachments were readily discernible on glass slides by naked eye at a concentration of 0.1 pg/mL antigen in PBS solution. The indirect detection of antigen using the clusters of immunogold nanoparticles is an efficient way to improve the detection sensitivity.