Microfluidic sensor using pH gradient with hybrid magnetoliposomes containing laccase immobilized nanocrystals.

A method using pH gradient to modify the phospholipid dissociation has been developed using hybrid magnetoliposomes with encapsulated enzyme laccase, which has been immobilized on hydrophilic magnetic nanocrystals (PAA-MNCs) retained in the reaction/detection zone of a microfluidic system. The hybrid magnetoliposomes act as micro containers, providing the safe transfer of the enzyme to the reaction/detection site, where it is retained, and preconcentration of the enzyme in this place occurs, which improves the system's sensitivity. The use of pH change involves the lack of external molecules to release the encapsulated materials.

An integrated microfluidic-based biosensor using a magnetically controlled MNPs-enzyme microreactor to determine cholesterol in serum with fluorometric detection.

This work provides a microfluidic-based biosensor to determine total cholesterol in serum based on integrating the reaction/detection zone of a microfluidic chip of a magnetically retained enzyme microreactor (MREµR) coupled with the remote fluorometric detection through a bifurcated fiber-optic bundle (BFOB) connected with a conventional spectrofluorometer. The method is based on developing the enzymatic hydrolysis and oxidation of cholesterol at microscale size using both enzymes (cholesterol esterase (ChE) and cholesterol oxidase (ChOx)) immobilized on magnetic nanoparticles (MNPs). The biocatalyst reactions were followed by monitoring the fluorescence decreasing by the naphtofluorescein (NF) oxidation in the presence of the previous HO formed.

Integrated microfluidic systems for fluorescence monitoring rapid kinetic reactions in bioanalysis.

A stopped-flow microfluidic fluorimetric biosensor to monitor alkaline phosphatase (ALP) activity and evaluate the potential inhibitors has been developed, integrating a magnetically retained enzyme microreactor (MREµR) in the reaction/detection zone of the microfluidic chip. The integration supposed the alignment of the MREµR at the sample compartment of a conventional spectrofluorometer using a 3D-printed device. The analytical signal is based on the fluorescence decrease in the signal obtained in the dephosphorylation reaction of the substrate 4-methylumbelliferone phosphate (4-MUP) by the retained ALP-MNPs in an alkaline medium caused by sulfonamides.

Study of the inhibition effects on glutathione peroxidase immobilized on MNPs using a stopped-flow microfluidic system.

A stopped-flow microfluidic system to monitor glutathione peroxidase (GPx) activity and evaluate potential inhibitors of the enzyme has been developed based on the integration of the microfluidic chip in the reaction/detection zone. This integration supposes the physical alignment at the optimal location of the microfluidic channel, both the magnetically retained enzyme microreactor (MREµR) and the remote luminescence detection using a focused bifurcated fiber optic bundle (BFOB) connected to a conventional spectrofluorometer detector. The method is based on the coupling of two competitive oxidative chemical reactions, in which glutathione (GSH) and homovanillic acid (HVA) competed for their interaction with hydrogen peroxide in the presence of the magnetically retained GPx-MNPs.

Usefulness of Hybrid Magnetoliposomes for Aminoglycoside Antibiotic Residues Determination in Food Using an Integrated Microfluidic System with Fluorometric Detection.

A new microfluidic approach using hybrid magnetoliposomes (h-MLs) containing hydrophobic magnetic nanoparticles (FeO@AuNPs-C12SH) and encapsulated -acetylcysteine has been developed in this research to determine aminoglycoside antibiotic (AAG) residues in food using -phthalaldehyde. Four AAGs, kanamycin, streptomycin, gentamicin, and neomycin, have been used as model analytes. The h-MLs have been used for reagent preconcentration and were retained using an external electromagnet device in the reaction/detection zone in a microfluidic system, inserted into the sample chamber of a conventional fluorimeter.

Development of an aptamer-based SPR-biosensor for the determination of kanamycin residues in foods.

A biosensor in which an affinity reaction occurs in the sensitive microzone through the use of specific aptamers to determine kanamycin residues in agri-food samples has been developed. It is an irreversible and continuous flow aptameric biosensor (aptasensor) in which the signal variations are monitored by surface plasmon resonance (SPR) measurements based on the specific interaction of the aptamer with the antibiotic. The signal variation is proportional to the analyte concentration.

Separation and characterization of liposomes using asymmetric flow field-flow fractionation with online multi-angle light scattering detection.

Liposomes, mainly formed by phospholipids and cholesterol that entrapped different compounds, were separated and characterized using asymmetric flow field-flow fractionation (AF4) coupled with a multi-angle light scattering detector (MALS). AF4 allows the separation of liposomes according to their hydrodynamic size, and the particle size can be estimated directly by their elution time. Besides, different synthesized liposome suspensions of liposomes with different species encapsulated in different places in liposomes were prepared with analytical purposes to be studied.

Luminescence continuous flow system for monitoring the efficiency of hybrid liposomes separation using multiphase density gradient centrifugation.

A method for monitoring the efficiency of the hybrid magnetoliposomes (h-MLs) separation using multiphase density gradient centrifugation (MDGC) coupled with a continuous flow system (CFS) is described. Several h-MLs suspensions containing hydrophobic magnetic gold nanoparticles (FeO@AuNPs-C12SH) and different fluorophores encapsulated have been synthesized using the rapid solvent evaporation (RSE) method. The MDGC system was prepared using a non-linear multiphase density gradient formed with a bottom layer with 100% (v/v) sucrose solution and six layers containing a mixture of sucrose solution (with concentrations ranged between 10 and 55% v/v), and fixed concentrations of ficoll (30% v/v) and percoll (15% v/v) solutions.

Integration of a microfluidic system into a conventional luminescence detector using a 3D printed alignment device.

A useful 3D printed device for the inside microfluidic integration into a conventional optical detector has been developed. The coupling system supposes the complete integration of a microfluidic device inside the sample compartment of a conventional spectrofluorimeter. For this purpose, a commercial chip-holder, including a microfluidic chip, was anchored inside the detector using a "lab-built" 3D printing alignment prototype.

Usefulness of magnetically-controlled MNPs-enzymes microreactors for the fluorimetric determination of total cholesterol in serum.

A new dynamic method containing a magnetically retained enzyme reactor (MRER) located in the reaction/detection zone of a flow injection (FI) system, has been used for the determination of total cholesterol in serum samples. The MRER was formed by a mixture ratio of 2/1 of immobilized enzymes cholesterol esterase (ChE) and cholesterol oxidase (COx) on magnetic nanoparticles (MNPs). The analytical signal is based on the fluorescence decreasing of the fluorophore naphtofluorescein (NF) due to its oxidation by the HO formed in the enzymatic reactions.

Automatic determination of coenzyme Q10 in food using cresyl violet encapsulated into magnetoliposomes.

A new type of magnetoliposomes (MLs), containing hydrophobic magnetic-gold nanoparticles and the long wavelength fluorophore cresyl violet, has been used for the determination of coenzyme Q10 (CoQ10). MLs were concentrated just before the detector, using a flow system and an external electromagnet device. The subsequent introduction of Triton X-100 and CoQ10 causes the MLs lysis and the cresyl violet oxidation, obtaining a decrease in the fluorescence signal.

Fluorometric determination of alkaline phosphatase activity in food using magnetoliposomes as on-flow microcontainer devices.

Liposomes containing magnetic gold nanoparticles (AuNPs) and an enzymatic substrate (4-methylumbelliferyl-phosphate) have been used as on-flow microcontainers for reagent preconcentration in a flow injection method for the determination of alkaline phosphatase (ALP) activity. The dynamic range of the calibration graph was 6.4 × 10(-3)-0.

Gold nanoparticle-biotinylated liposome hybrids as analytical reagents for biotin determination using a competitive assay and resonance light scattering detection.

The preparation of hybrid nanostructures formed by gold nanoparticles (AuNPs) into biotinylated liposomes and their analytical application are presented. The surface of negatively charged AuNPs was modified with 1-dodecanethiol and the NPs were encapsulated into biotinylated liposomes using the rapid solvent evaporation method. Liposomes were resized by both mechanical shaking and ultrasound treatments and filled liposomes were separated from empty liposomes using sucrose density gradient centrifugation.

Luminescent determination of flavonoids in orange juices by LC with post-column derivatization with aluminum and terbium.

A new post-column derivatization system is described and applied to the determination of flavonoids in citric beverages after their separation by LC using a monolithic column. The derivatization involves the formation of the chelates of the analytes with aluminum (III) and terbium (III) in the presence of the surfactant SDS and the measurement of the terbium sensitized luminescence at lambda(ex) 360 and lambda(em) 545 nm. Naringin, hesperidin, quercetin, naringenin, and kaempferol have been chosen as analyte models.

High throughput bioassays using nanoparticles.

An overview of the usefulness of different nanoparticles to improve the features of high throughput separation and individual and multiplexed detection bioassays is presented. Although the development of microarray and microfluidic systems has expanded the capabilities of these high throughput assays, the combined use of NPs and these devices has provided them with new applications in drug discovery, proteomic and genomic studies, and clinical diagnosis. This article reviews the wide application field of magnetic, gold, silver, semiconductor and other nanoparticles in high throughput bioassays.

Determination of fluoroquinolones in milk samples by postcolumn derivatization liquid chromatography with luminescence detection.

A liquid chromatography (LC) method with luminescence detection for the determination of eight quinolone antibiotics is reported. The system encompasses three consecutive steps: (a) chromatographic separation using reverse-phase mode (RP-LC), (b) postcolumn derivatization reaction, and (c) luminescence detection by monitoring fluorescence (FL) and time-resolved (TR) signals. The derivatization step is based on the reaction between quinolones and terbium(III) to form luminescent chelates, which were determined at lambda(ex) 340 and lambda(em) 545 nm (FL mode) or at lambda(ex) 281 and lambda(em) 545 nm (TR mode).