Live synthesis of selective carbon dots as fluorescent probes for cobalt determination in water with an automatic microanalyzer.

A new strategy integrating the straight synthesis of carbon dots (CDs) and their direct use for the determination of heavy metals by means of fluorescence quenching is presented. The proposal consists of a modular analyzer, which includes a low temperature co-fired ceramics (LTCC) microreactor for the synthesis of CDs and a cyclic olefin copolymer (COC) microfluidic platform, which automatically performs a reverse flow injection analysis (rFIA) protocol for the determination of heavy metal ions in water by CD fluorescence quenching. As a proof of concept, nitrogen-doped CDs were synthesized from acrylic acid and ethylenediamine (ED) with quantum yields (QYs) of up to 44%, which are selective to cobalt. With the described system, we synthesized homogeneous CDs without the need for further purification and with the minimum consumption of reagents, and optimized fluorescence measurements can be performed with freshly obtained luminescent nanomaterials that have not undergone decomposition processes. They have an average hydrodynamic diameter of 4.2 ± 0.9 nm and maximum excitation and emission wavelengths at 358 nm and 452 nm, respectively. The system allows the automatic dilution and buffering of the synthesized CDs and the sample prior to the determination of cobalt. The concentration of cobalt was determined with good sensitivity and a limit of detection of 7 µg·L-1 with a linear range of 0.02-1 mg·L-1 of Co2+. Spiked tap water and river water samples were analyzed, obtaining recovery from 98 to 104%. This demonstrates the potential of the equipment as an efficient on-site control system for heavy metal monitoring in water.

Microanalyser Prototype for On-Line Monitoring of Copper(II) Ion in Mining Industrial Processes.

A microanalyzer prototype for copper(II) ion monitoring in mining industrial processes is presented. The microanalyzer is designed as an assembly of different modules, each module being responsible for a unit operation. In order to optimize the industrial processes, the microanalyzer can automate all sample management, signal processing, and mathematical calculations and wirelessly transfer data to a control room. The determination of copper(II) ion is done using a colorimetric reaction and the microanalyser performs autocalibration by in situ dilution of a stock solution, matching the higher analyte concentration of the working range defined for the sample to be determined, using a multicommutation approach. The performance of the microanalyzer for monitoring copper(II) ion in water effluents of mining facilities was optimized in the working range from 1 to 10 mg/L to match Mexican environmental law regulations, which allow a maximum concentration of 4 mg/L of copper(II) ion in these circumstances.

Microfabrication of Monolithic Microfluidic Platforms Using Low Temperature Co-Fired Ceramics Suitable for Fluorescence Imaging.

In this work, the influence of laser ablation and lamination parameters in the fabrication of embedded microstructures using Low Temperature Co-Fired Ceramics, have been studied. First, the influence of laser ablation parameters in the dimensions of fabricated microchannels in Low Temperature Co-Fired Ceramics substrates was characterized and strategies for tailoring the microchannels aspect ratios are described. The influence of lamination conditions on the fabrication of monolithically embedded microstructures is presented. Thereafter, a ceramic microfluidic platform, was constructed using a multilayer approach. The ceramic microfluidic platforms incorporate three independent inlet channels and a microfluidic chamber with an monolithically integrated transparent optical windows. The construction procedure used ensures monolithic ceramic devices with homogeneous surface chemistry as well as homogeneous physical properties. Fluorescence dyes were used in order to characterize the hydrodynamic focusing as a function of flow rate ratio of the microfluidic chamber inlets. The results obtained open the possibility of studying chemical process, in static or flow conditions using fluorescence imaging, within the traditional fields of LTCC technology, such as high-temperature or organic solvents applications, while using a simple fabrication procedure suitable for low cost mass production.

Versatile lock and key assembly for optical measurements with microfluidic platforms and cartridges.

A novel and versatile optical reader for microfluidic platforms is presented. The reader includes a modular insertion port based on the lock and key concept for reproducible alignment with a miniaturized optical detection system comprising an interchangeable light emitting diode (LED) and a photodiode. The modular nature of the insertion port allows the use of microfluidic platforms in variable shapes and fluidic configurations. Three different analytical methodologies based on absorbance or fluorescence measurements were used to demonstrate the flexibility and reproducibility of the proposed experimental setup.

Magnetic actuator for the control and mixing of magnetic bead-based reactions on-chip.

While magnetic bead (MB)-based bioassays have been implemented in integrated devices, their handling on-chip is normally either not optimal--i.e. only trapping is achieved, with aggregation of the beads--or requires complex actuator systems. Herein, we describe a simple and low-cost magnetic actuator to trap and move MBs within a microfluidic chamber in order to enhance the mixing of a MB-based reaction. The magnetic actuator consists of a CD-shaped plastic unit with an arrangement of embedded magnets which, when rotating, generate the mixing. The magnetic actuator has been used to enhance the amplification reaction of an enzyme-linked fluorescence immunoassay to detect Escherichia coli O157:H7 whole cells, an enterohemorrhagic strain, which have caused several outbreaks in food and water samples. A 2.7-fold sensitivity enhancement was attained with a detection limit of 603 colony-forming units (CFU) /mL, when employing the magnetic actuator.

Fluorescence Imaging Characterization of the Separation Process in a Monolithic Microfluidic Free-Flow Electrophoresis Device Fabricated Using Low-Temperature Co-Fired Ceramics.

A monolithic microfluidic free-flow electrophoresis device, fabricated using low-temperature co-fired ceramic technology, is presented. The device integrates gold electrodes and a 20 µm thick transparent ceramic optical window, suitable for fluorescence imaging, into a multilevel microfluidic chamber design. The microfluidic chamber consists of a 60 µm deep separation chamber and two, 50 µm deep electrode chambers separated by 10 µm deep side channel arrays. Fluorescence imaging was used for in-chip, spatial-temporal characterization of local pH variations in separation conditions as well as to characterize the separation process. The device allowed baseline resolution separation of a sample mixture of Fluorescein, Rhodamine 6G, and 4-Methylumbelliferone at pH 7.0, in only 6 s, using 378 V.s/cm. The results demonstrate the possibility of studying a chemical process using fluorescence imaging within the traditional fields of low-temperature co-fired ceramics technology, such as high-electrical-field applications, while using a simple fabrication procedure suitable for low-cost mass production.

Biomedical point-of-care microanalyzer for potentiometric determination of ammonium ion in plasma and whole blood.

Some inborn errors of metabolism and other diseases can result in increasing blood ammonium (hyperammonemia episodes), which can cause serious neurological complications in patients or even death. Early diagnosis, follow up and treatment are essential to minimize irreversible damages in brain. Currently, adequate analytical instrumentation for the necessary ammonium bedside determination is not available in all health centers but only in clinical laboratories of reference hospitals. We therefore have developed a low cost and portable potentiometric Point-of-Care microanalyzer (POC) to address this problem. It consists of a cyclic olefin copolymer-based microanalyzer, the size of a credit card and working in continuous flow, which integrates microfluidics, a gas-diffusion module and a potentiometric detection system. The analytical features achieved are a linear range from 30 to 1000 µmol L-1 NH4+, a detection limit of 18 µmol L-1 NH4+ and a required sample volume of 100 µL, which comply with the medical requirements. Plasma and blood samples are analyzed with no significant differences observed between ammonium concentrations obtained with both the proposed microanalyzer and the reference method. This demonstrates the value of the developed POC for bedside clinical applications.

Soluble reactive phosphorous determination in wastewater treatment plants by automatic microanalyzers.

The analysis of soluble reactive phosphate (SRP) in water is key to control water quality. In order to continuous monitor orthophosphate content in water during treatment processes and in the effluents of wastewater treatment plants, conventional procedures, usually performed in a laboratory, must be adapted. This means pursuing efforts on miniaturizing systems to operate in situ and automating analytical methods to work on-line. The design, construction and evaluation of an automatic and low cost cyclic olefin copolymer (COC)-based spectrophotometric microanalyzer, capable of operating in unattended conditions, is presented to monitor soluble reactive phosphorous, as orthophosphate ion, in wastewater samples coming from sewage treatment plants. The microsystem, constructed by CNC micromilling and using a multilayer approach, integrates microfluidics to carry out the phosphomolybdenum blue (PMB) reaction and an optical flow-cell for the spectrophotometric orthophosphate determination in a single polymeric substrate smaller than a credit card. It is connected to a compact optical detection system composed by a LED emitting at 660 nm and a PIN-photodiode, both integrated in a PCB. Flow management is automatically performed by programmed microvalves and micropumps, which control autocalibration processes and allow unattended operation. Analytical features after the optimization of the microfluidic platform and the chemical and the hydrodynamic variables, were a linear range from 0.09 to 32 mg L-1 P and a detection limit of 0.03 mg L-1 P with a sampling rate of 24 samples h-1, demonstrating the microanalyzer suitability for SRP monitoring in water. Moreover, real samples were analyzed obtaining promising results.

Monitoring of total potassium in winemaking processes using a potentiometric analytical microsystem.

Innovation in products and processes, traceability, food security and quality control are inherent challenges in agri-food sector. Trends in wine production are focused on obtaining natural wines with less chemical intervention. Following this goal, a low cost miniaturized, easy-to-use and highly automated microanalyzer to monitor total potassium in winemaking processes is presented. The microsystem monolithically integrates microfluidics as well as a potentiometric detection system and does not require any sample pretreatment. The analytical features provided are a linear range from 250 to 4000 mg L-1 K+, covering all the concentrations expected in must and wine samples, a detection limit of 75 ± 12 mg L-1 K+, and an adequate reproducibility and repeatability. Sample throughput is calculated at 20 h-1 with a waste volume generation lower than 4 mL per analysis. The microsystem lifetime is at least 4 months. Different wine and grape juice samples have been analyzed reaching outstanding results.

A monolithic continuous-flow microanalyzer with amperometric detection based on the green tape technology.

The development of micro total analysis systems (muTAS) has become a growing research field. Devices that include not only the fluidics and the detection system but also the associated electronics are reported scarcely in the literature because of the complexity and the cost involved for their monolithic integration. Frequently, dedicated devices aimed at solving specific analytical problems are needed. In these cases, low-volume production processes are a better alternative to mass production technologies such as silicon and glass. In this work, the design, fabrication, and evaluation of a continuous-flow amperometric microanalyzer based on the green tape technology is presented. The device includes the microfluidics, a complete amperometric detection system, and the associated electronics. The operational lifetime of the working electrode constitutes a major weak point in electrochemical detection systems, especially when it is integrated in monolithic analytical devices. To increase the overall system reliability and its versatility, it was integrated following an exchangeable configuration. Using this approach, working electrodes can be readily exchanged, according to the analyte to be determined or when their surfaces become passivated or poisoned. Furthermore, the electronics of the system allow applying different voltamperometric techniques and provide four operational working ranges (125, 12.5, 1.25, and 0.375 microA) to do precise determinations at different levels of current intensity.

Automated analytical microsystem for the spectrophotometric monitoring of titratable acidity in white, rosé and red wines.

The design, construction and evaluation of a low-cost cyclic olefin copolymer (COC)-based continuous flow microanalyzer with optical detection to determine the titratable acidity content of wine is here presented. The analysis method is based on the monitoring of the blue coloration decrease of a buffered bromothymol blue (BTB) solution in the presence of the acidic compounds of wine. The microanalyzer monolithically integrates the required microfluidic motifs as well as an optical flow cell where the measurements are performed by using a miniaturized and versatile photometric detection system. Fluid management is totally automated by the use of computer-controlled microvalves, permitting the automatic calibration of the system as well as the automatic sampling, including in-line dilution and analysis. The reduced size of the whole system along with its high simplicity and automation make it suitable for its application to the on-line monitoring of titratable acidity during wine-making processes. With the optimal conditions, a linear range up to 0.50 g L-1 tartaric acid, a quantification limit (LOQ) of 0.01 g L-1 and a detection limit (LOD) of 0.004 g L-1 were obtained, covering the most common acidity content of musts and wines. A sampling rate up to 26 h-1 could be achieved, consuming less than 3 mL of inexpensive reagents per analysis and requiring no pretreatment of the sample. The microsystem has been successfully applied to the quantification of the titratable acidity content of several wine samples, being the results in excellent agreement with the ones obtained by the reference method.

Highly integrated autonomous lab-on-a-chip device for on-line and in situ determination of environmental chemical parameters.

The successful integration of sample pretreatment stages, sensors, actuators and electronics in microfluidic devices enables the attainment of complete micro total analysis systems, also known as lab-on-a-chip devices. In this work, we present a novel monolithic autonomous microanalyzer that integrates microfluidics, electronics, a highly sensitive photometric detection system and a sample pretreatment stage consisting on an embedded microcolumn, all in the same device, for on-line determination of relevant environmental parameters. The microcolumn can be filled/emptied with any resin or powder substrate whenever required, paving the way for its application to several analytical processes: separation, pre-concentration or ionic-exchange. To promote its autonomous operation, avoiding issues caused by bubbles in photometric detection systems, an efficient monolithic bubble removal structure was also integrated. To demonstrate its feasibility, the microanalyzer was successfully used to determine nitrate and nitrite in continuous flow conditions, providing real time and continuous information.

Gold/silver/gold trilayer films on nanostructured polycarbonate substrates for direct and label-free nanoplasmonic biosensing.

Ultrasmooth gold/silver/gold trilayer nanostructured plasmonic sensors were obtained using commercial Blu-ray optical discs as nanoslits-based flexible polymer substrates. A thin gold film was used as an adhesion and nucleation layer to improve the chemical stability and reduce the surface roughness of the overlying silver film, without increasing ohmic plasmon losses. The structures were physically and optically characterized and compared with nanostructures of single gold layer. Ultrasmooth and chemically stable trilayer nanostructures with a surface roughness <0.5 nm were obtained following a simple and reproducible fabrication process. They showed a figure of merit (FOM) value up to 69.2 RIU-1 which is significantly higher (more than 95%) than the gold monolayer counterpart. Their potential for biosensing was demonstrated by employing the trilayer sensor for the direct and refractometric (label-free) detection of C-reactive protein (CRP) biomarker in undiluted urine achieving a Limit of Detection (LOD) in the pM order.

Multi-parametric polymer-based potentiometric analytical microsystem for future manned space missions.

The construction and evaluation of a Cyclic Olefin Copolymer (COC)-based continuous flow potentiometric microanalyzer to simultaneously monitor potassium, chloride and nitrate ions in samples from an on-board water recycling process expected to be installed in future manned space missions is presented. The main goals accomplished in this work address the specific required characteristics for a miniaturized on-line monitoring system to control water quality in such missions. To begin with, the integration of three ion-selective electrodes (ISEs) and a reference electrode in a compact microfluidic platform that incorporates a simple automatic autocalibration process allows obtaining information about the concentration of the three ions with optimal analytical response characteristics, but moreover with low reagents consumption and therefore with few waste generation, which is critical for this specific application. By a simple signal processing (signal removal) the chloride ion interference on the nitrate electrode response can be eliminated. Furthermore, all fluidics management is performed by computer-controlled microvalves and micropumps, so no manual intervention of the crew is necessary. The analytical features provided by the microsystem after the optimization process were a linear range from 6.3 to 630 mg L-1 and a detection limit of 0.51 mg L-1 for the potassium electrode, a linear range from 10 to 1000 mg L-1 and a detection limit of 1.58 mg L-1 for the chloride electrode and a linear range from 10 to 1000 mg L-1 and a detection limit of 3.37 mg L-1 for the nitrate electrode with a reproducibility (RSD) of 4%, 2% and 3% respectively. Sample throughput was 12 h-1 with a reagent consumptions lower than 2 mL per analysis.

Rapid Prototyping of a Cyclic Olefin Copolymer Microfluidic Device for Automated Oocyte Culturing.

Assisted reproductive technology (ART) can benefit from the features of microfluidic technologies, such as the automation of time-consuming labor-intensive procedures, the possibility to mimic in vivo environments, and the miniaturization of the required equipment. To date, most of the proposed approaches are based on polydimethylsiloxane (PDMS) as platform substrate material due to its widespread use in academia, despite certain disadvantages, such as the elevated cost of mass production. Herein, we present a rapid fabrication process for a cyclic olefin copolymer (COC) monolithic microfluidic device combining hot embossing-using a low-temperature cofired ceramic (LTCC) master-and micromilling. The microfluidic device was suitable for trapping and maturation of bovine oocytes, which were further studied to determine their ability to be fertilized. Furthermore, another COC microfluidic device was fabricated to store sperm and assess its quality parameters over time. The study herein presented demonstrates a good biocompatibility of the COC when working with gametes, and it exhibits certain advantages, such as the nonabsorption of small molecules, gas impermeability, and low fabrication costs, all at the prototyping and mass production scale, thus taking a step further toward fully automated microfluidic devices in ART.

Rapid Prototyping of a Cyclic Olefin Copolymer Microfluidic Device for Automated Oocyte Culturing.

Assisted reproductive technology (ART) can benefit from the features of microfluidic technologies, such as the automation of time-consuming labor-intensive procedures, the possibility to mimic in vivo environments, and the miniaturization of the required equipment. To date, most of the proposed approaches are based on polydimethylsiloxane (PDMS) as platform substrate material due to its widespread use in academia, despite certain disadvantages, such as the elevated cost of mass production. Herein, we present a rapid fabrication process for a cyclic olefin copolymer (COC) monolithic microfluidic device combining hot embossing-using a low-temperature cofired ceramic (LTCC) master-and micromilling. The microfluidic device was suitable for trapping and maturation of bovine oocytes, which were further studied to determine their ability to be fertilized. Furthermore, another COC microfluidic device was fabricated to store sperm and assess its quality parameters over time. The study herein presented demonstrates a good biocompatibility of the COC when working with gametes, and it exhibits certain advantages, such as the nonabsorption of small molecules, gas impermeability, and low fabrication costs, all at the prototyping and mass production scale, thus taking a step further toward fully automated microfluidic devices in ART.

Low cost and compact analytical microsystem for carbon dioxide determination in production processes of wine and beer.

The design, construction and evaluation of a low cost, cyclic olefin copolymer (COC)-based continuous flow microanalyzer, with optical detection, to monitor carbon dioxide in bottled wines and beers as well as in fermentation processes, is presented. The microsystem, constructed by computer numerically controlled (CNC) micromilling and using a multilayer approach, integrates microfluidics, gas-diffusion module and an optical flow-cell in a single polymeric substrate. Its size is slightly bigger than a credit card, exactly 45 × 60 × 4 mm in the microfluidic and diffusion module zone and 22.5 × 40 × 3 mm in the flow-cell zone. The gas-diffusion module is based on a hydrophobic polyvinylidene fluoride (PVDF) membrane, which allows the transfer of the carbon dioxide present in the sample to a bromothymol blue (BTB) pH-sensitive acceptor solution, where the color change is measured optically. The detection system consisted of a LED with an emission peak at 607 nm and a photodiode integrated in a printed circuit board (PCB). The obtained analytical features after the optimization of the microfluidic platform and hydrodynamic variables are a linear range from 255 to 10000 mg L(-1) of CO2 and a detection limit of 83 mg L(-1) with a sampling rate of 30 samples h(-1).

Potentiometric analytical microsystem based on the integration of a gas-diffusion step for on-line ammonium determination in water recycling processes in manned space missions.

The design, construction and evaluation of a versatile cyclic olefin copolymer (COC)-based continuous flow potentiometric microanalyzer to monitor the presence of ammonium ion in recycling water processes for future manned space missions is presented. The microsystem integrates microfluidics, a gas-diffusion module and a detection system in a single substrate. The gas-diffusion module was integrated by a hydrophobic polyvinylidene fluoride (PVDF) membrane. The potentiometric detection system is based on an all-solid state ammonium selective electrode and a screen-printed Ag/AgCl reference electrode. The analytical features provided by the analytical microsystem after the optimization process were a linear range from 0.15 to 500 mg L(-1) and a detection limit of 0.07 ± 0.01 mg L(-1). Nevertheless, the operational features can be easily adapted to other applications through the modification of the hydrodynamic variables of the microfluidic platform.

Gas diffusion as a new fluidic unit operation for centrifugal microfluidic platforms.

A centrifugal microfluidic platform prototype with an integrated membrane for gas diffusion is presented for the first time. The centrifugal platform allows multiple and parallel analysis on a single disk and integrates at least ten independent microfluidic subunits, which allow both calibration and sample determination. It is constructed with a polymeric substrate material and it is designed to perform colorimetric determinations by the use of a simple miniaturized optical detection system. The determination of three different analytes, sulfur dioxide, nitrite and carbon dioxide, is carried out as a proof of concept of a versatile microfluidic system for the determination of analytes which involve a gas diffusion separation step during the analytical procedure.

Biparametric potentiometric analytical microsystem for nitrate and potassium monitoring in water recycling processes for manned space missions.

The construction and evaluation of a Low Temperature Co-fired Ceramics (LTCC)-based continuous flow potentiometric microanalyzer prototype to simultaneously monitor the presence of two ions (potassium and nitrate) in samples from the water recycling process for future manned space missions is presented. The microsystem integrates microfluidics and the detection system in a single substrate and it is smaller than a credit card. The detection system is based on two ion-selective electrodes (ISEs), which are built using all-solid state nitrate and potassium polymeric membranes, and a screen-printed Ag/AgCl reference electrode. The obtained analytical features after the optimization of the microfluidic design and hydrodynamics are a linear range from 10 to 1000 mg L(-1) and from 1.9 to 155 mg L(-1) and a detection limit of 9.56 mg L(-1) and 0.81 mg L(-1) for nitrate and potassium ions respectively.