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The ongoing coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has led to a global public health disaster. The current gold standard for the diagnosis of infected patients is real-time reverse transcription-quantitative PCR (RT-qPCR). As effective as this method may be, it is subject to false-negative and -positive results, affecting its precision, especially for the detection of low viral loads in samples. In contrast, digital PCR (dPCR), the third generation of PCR, has been shown to be more effective than the gold standard, RT-qPCR, in detecting low viral loads in samples. In this review article, we selected publications to show the broad-spectrum applications of dPCR, including the development of assays and reference standards, environmental monitoring, mutation detection, and clinical diagnosis of SARS-CoV-2, while comparing it analytically to the gold standard, RT-qPCR. In summary, it is evident that the specificity, sensitivity, reproducibility, and detection limits of RT-dPCR are generally unaffected by common factors that may affect RT-qPCR. As this is the first time that dPCR is being tested in an outbreak of such a magnitude, knowledge of its applications will help chart a course for future diagnosis and monitoring of infectious disease outbreaks.

The digital polymerase chain reaction (dPCR) is an irreplaceable variant of PCR techniques due to its capacity for absolute quantification and detection of rare deoxyribonucleic acid (DNA) sequences in clinical samples. Image processing methods, including micro-chamber positioning and fluorescence analysis, determine the reliability of the dPCR results. However, typical methods demand high requirements for the chip structure, chip filling, and light intensity uniformity. This research developed an image-to-answer algorithm with single fluorescence image capture and known image-related error removal. We applied the Hough transform to identify partitions in the images of dPCR chips, the 2D Fourier transform to rotate the image, and the 3D projection transformation to locate and correct the positions of all partitions. We then calculated each partition's average fluorescence amplitudes and generated a 3D fluorescence intensity distribution map of the image. We subsequently corrected the fluorescence non-uniformity between partitions based on the map and achieved statistical results of partition fluorescence intensities. We validated the proposed algorithms using different contents of the target DNA. The proposed algorithm is independent of the dPCR chip structure damage and light intensity non-uniformity. It also provides a reliable alternative to analyze the results of chip-based dPCR systems.

A microfluidic-based digital polymerase chain reaction (dPCR) chip requires precise temperature control as well as uniform temperature distribution to ensure PCR efficiency. However, measuring local temperature and its distribution over thousands of L/nL-volume samples with minimum disturbance is challenging. Here, we present a method of non-contact localized temperature measurement for determination of the non-uniformity of temperature distribution over a dPCR chip. We filled the dPCR chip with a PCR solution containing amplified DNA fragments with a known melting temperature (TM). We then captured fluorescent images of the chip when it was heated from 70 to 99 °C, plotted the fluorescence intensity of each partition as a function of temperature, and calculated measured TM values from each partition. Finally, we created a 3-D map of the dPCR chip with the measured TM as the parameter. Even when the actual TM of the PCR solution was constant, the measured TM value varied between locations due to temperature non-uniformity in the dPCR chip. The method described here thereby characterized the distribution of temperature non-uniformity using a PCR solution with known TM as a temperature sensor. Among the non-contact temperature measurement methods, the proposed TM-based method can determine the temperature distribution within the chip, instead of only at the chip surface. The method also does not suffer from the undesirable photobleaching effect of fluorescein-based temperature measurement method. Temperature determination over the dPCR chip based on TM allowed us to calibrate the temperature sensor and improve the dPCR configuration and precision. This method is also suitable for determining the temperature uniformity of other microarray systems where there is no physical access to the system and thus direct temperature measurement is not possible.

Currently, the ongoing pandemic has created an unprecedented pressure for the development of various analytical assays and especially their immediate practical applications. A number of tests based on different principles such as immunoassay [1], electrochemistry [2], nucleic acid amplification tests (NAATs) [3] and others have been reported [4]. Immunoassays and NAATs are currently being utilized clinically, while systems based on several other principles did not go further than the acceptance of manuscripts for publication.

The use of microelectrodes or arrays of microelectrodes is beneficial for electrochemical systems with minute volumes such as sensors embedded in microfluidics. Here, we present an approach to fabricate nanostructured gold amalgam (AuxHgy) via a single-step co-deposition method. The gold amalgam was selectively deposited through a lithography mask with opening areas of diameter 4 μm, forming an array of microelectrodes with a nanostructured surface. This surface was three orders of magnitude larger than that defined by lithography. The surface material properties were modified by the subsequent removal of Hg using a dealloying method in order to enable operation in the positive potential region similar to conventional Au electrodes. We performed electrochemiluminescence (ECL) with an entire array in an electrochemical cell and observed the ECL from a single microelectrode to study details of the ECL reactions and their kinetics. The nanostructured AuxHgy microelectrodes have overvoltage to hydrogen evolution, thus allowing the electrodes to operate in more negative potentials compared with noble metals such as Au and Pt, resulting in an ultrasensitive detection of materials such as DNA and proteins.

We proposed and demonstrated a preparation method of (001) preferentially oriented stress-free AlN piezoelectric thin films. The AlN thin films were deposited by a reactive sputtering technique at substrate temperatures up to 330 °C using a dual Kaufman ion-beam source setup. We deposited the AlN on Si (100), Si (111), amorphous SiO2, and a (001) preferentially oriented Ti thin film and compared their crystallographic, optical, and piezoelectric properties. The AlN thin films deposited on the (001) preferentially oriented Ti thin films have the highest crystallographic quality. The stress-free AlN reached a high value of the piezoelectric coefficient d33 = (7.33 ± 0.08) pC·N−1. The properties of the AlN thin film prepared at such low temperatures are suitable for numerous microelectromechanical systems, piezoelectric sensors, and actuators monolithically integrated with complementary metal-oxide-semiconductor signal-processing circuits.

Microcalorimetry has been widely used to measure and characterize the heat change in phase transition, resolution mixing, and chemical/biochemical reactions. In this review, we discuss progress in the development of high-performance microfabricated calorimeters starting with their design, fluid handling, and temperature measurement. The considerations of chip design are highlighted. Then, a few representative applications are introduced and their chip parameters and properties are compared. Finally, trends and prospects of microcalorimeters for high-performance applications, such as enthalpy measurements in minute volumes of chemicals suitable for drug discovery as well as bio-applications in energy balance monitoring of living cells and investigation of their thermogenesis, are discussed.

Microfluidic technology is revolutionising a number of scientific fields, including chemistry, biology, diagnostics, and engineering. The ability to manipulate fluids and objects within networks of micrometre-scale channels allows reductions in processing and analysis times, reagent and sample consumption, and waste production, whilst allowing fine control and monitoring of chemical or biological processes. The integration of multiple components and processes enable “lab-on-a-chip” devices and “micro total analysis systems” that have applications ranging from analytical chemistry, organic synthesis, and clinical diagnostics to cell biology and tissue engineering. This concise, easy-to-read book is perfectly suited for instructing newcomers on the most relevant and important aspects of this exciting and dynamic field, particularly undergraduate and postgraduate students embarking on new studies, or for those simply interested in learning about this widely applicable technology. Written by a team with more than 20 years of experience in microfluidics research and teaching, the book covers a range of topics and techniques including fundamentals (e.g. scaling laws and flow effects), microfabrication and materials, standard operations (e.g. flow control, detection methods) and applications. Furthermore, it includes questions and answers that provide for the needs of students and teachers in the area.

PCR has become one of the most valuable techniques currently used in bioscience, diagnostics and forensic science. Here we review the history of PCR development and the technologies that have evolved from the original PCR method. Currently, there are two main areas of PCR utilization in bioscience: high-throughput PCR systems and microfluidics-based PCR devices for point-of-care (POC) applications. We also discuss the commercialization of these techniques and conclude with a look into their modifications and use in innovative areas of biomedicine. For example, real-time reverse transcription PCR is the gold standard for SARS-CoV-2 diagnoses. It could also be used for POC applications, being a key component of the sample-to-answer system.

We propose and demonstrate an unconventional method suitable for releasing microelectromechanical systems devices containing an Al layer by wet etching using SiO2 as a sacrificial layer. We used 48% HF solution in combination with 20% oleum to keep the HF solution water-free and thus to prevent attack of the Al layer, achieving an outstanding etch rate of thermally grown SiO2 of &asymp, 1 &micro, m&middot, min&minus, 1. We also verified that this etching solution only minimally affected the Al layer, as the chip immersion for &asymp, 9 min increased the Al layer sheet resistance by only &asymp, 7.6%. The proposed etching method was performed in an ordinary fume hood in a polytetrafluorethylene beaker at elevated temperature of &asymp, 70 &deg, C using water bath on a hotplate. It allowed removal of the SiO2 sacrificial layer in the presence of Al without the necessity of handling highly toxic HF gas.

We introduced a microfluidic device for high-efficiency single cell capture and their subsequent extraction. It was fabricated by using a deep reactive ion etching process taking advantage of the lag effect. We accomplished single cell capture based on the least flow resistance path principle as well as independent extraction of individual captured cells to perform their subsequent analysis. The device was performed with >99% single cell capture efficiency.

Determination of microbolometer thermal properties such as thermal capacitance, conductance, time constant, and IR responsivity is of the utmost importance as they directly influence microbolometer performance. Here we show a technique to measure them by using a minimized self-heating effect, thus leading to their precise determination via measurements based on an AC-biased Wheatstone bridge containing a microbolometer. The bridge outputs were subtracted from each other by a differential voltage preamplifier with its output processed by a lock-in amplifier. The lock-in amplifier output as a function of the amplitude of AC bias provided an amplitude of microbolometer thermal conductivity. A microbolometer temperature response to pulse irradiation of its membrane provided the value of its thermal time constant and, thus, its thermal capacitance. Finally, we also extracted microbolometer responsivity using a blackbody IR source. The method was experimentally verified using a micromachined bolometer, which showed excellent agreement with the analytical solution.

Graphene has been of immense interest for its interesting electronic properties, such as being a zero-band gap semiconductor. However, to be able to usefully employ graphene for electronics and electronic-transduction system sensors and biosensors, one needs to open this band gap. This proofs to be challenging on reproducible, scalable way. There are other 2D monoelemental materials that exhibit useful band gap and which can be used for field effect transistor- (FET-) based sensing and biosensing. Here we discuss trends in the development of FET-based sensors utilizing 2D phosphorene, arsenene, antimonene, silicene, and germanene.

Carbon nanotubes (CNTs) have attracted significant attention due to their remarkable mechanical and electrical properties. Although it is assumed that the most important questions about CNTs have been addressed, the opposite is true. CNTs have high mechanical stiffness and electrical conductivity and, due to their small diameter and size, the measurement of those properties at nanoscale level is challenging. Here, we present a unique method to determine their mechanical and electrical properties by using interactions between vertically aligned multiwall CNTs and a metal-coated tipless atomic force microscope cantilever. We used a force–distance measurement (FDM) method with concurrent monitoring of electrical current. We could identify the number of CNTs in contact with the cantilever, stiffness of each individual CNT, the adhesion force between the cantilever and individual CNTs, and the electrical conductivity of each CNT. The method is also suitable for characterizing other vertically aligned nanostructured materials, such as nanowires. This method addresses the long-standing problem of property determination of materials such as CNTs and nanowires and is an important addition to the toolbox of nanoscale characterization methods.

Availability of potable water is a problem especially in developing countries as their water sources are often contaminated biologically as well as by heavy metals. Electrochemical methods are suitable for field application to monitor heavy metal contents. Noble metal electrodes with large surface areas are the subject of intensive development as they can yield an improved signal-to-noise ratio, enhanced sensitivity, and lower limits of detection (LOD). Here, we present a nanostructured Au microelectrode array subsequently modified by selective electrodeposition in lithographically defined circles within a partially cross-linked gelatin layer. This method increased its surface area by a factor of ≈1440 in comparison with an original lithography-based prepared array. The Au surface properties can be tailored by a degree of gelatin layer cross-linking. We used this array for an ultrasensitive detection of the As3+ ions content by stripping voltammetry achieving LOD of ≈0.0212 parts per billion (signal-to-no...

Microcalorimetry has been widely used in measuring cellular metabolic heat to study bioprocesses, such as metabolism. However, it is still limited by insufficient sensitivity and system complexity, especially for the direct measurements of individual cells. Here, we present a droplet-based simple differential microcalorimetric system for determining the real-time energy balance of the ciliate protist, Paramecium caudatum. We utilized the platform to dynamically monitor its defensive behavior and measured the temperature change. Then we used heat balance equation and calculated corresponding dissipated power and energy with an ultimate resolution of ≈14 nW. The results showed that the defensive behavior by the ejection of the trichocysts’ content consumed energy of ≈0.75 mJ per cell, and the dissipated power was ≈303.8 μW. This differential microcalorimetric platform can be used to study the metabolic heat of protist metabolism as well as other individual cells, helping us to understand this widespread, yet little-known, biological phenomenon from a new perspective.

The digital polymerase chain reaction (dPCR) multiplexing method can simultaneously detect and quantify closely related deoxyribonucleic acid sequences in complex mixtures. The dPCR concept is continuously improved by the development of microfluidics and micro- and nanofabrication, and different complex techniques are introduced. In this review, we introduce dPCR techniques based on sample compartmentalization, droplet- and chip-based systems, and their combinations. We then discuss dPCR multiplexing methods in both laboratory research settings and advanced or routine clinical applications. We focus on their strengths and weaknesses with regard to the character of biological samples and to the required precision of such analysis, as well as showing recently published work based on those methods. Finally, we envisage possible future achievements in this field.

We proposed a method to control and minimize residual stress in [001] preferentially oriented Ti thin films deposited by a Kaufman ion-beam source using a substrate temperature during deposition (T) as the parameter. We determined the residual stress, corresponding lattice parameters, and thickness of deposited films using X-ray diffraction and X-ray reflectivity measurements. We showed that the Ti film deposited at T ≈ 273 °C was stress-free with corresponding lattice parameters a0 and c0 of (2.954 ± 0.003) A and (4.695 ± 0.001) A, respectively. The stress-free sample has the superior crystallographic quality and pure [001] orientation. The Ti thin films were oriented with the c-axis parallel to the surface normal. We also investigated root mean square of surface roughness of deposited films by atomic force microscopy and it was in the range from ≈ 0.58 nm to ≈ 0.71 nm. Such smooth and stress-free layers are suitable for microelectromechanical systems.

We present a portable lock-in amplifier-based electrochemical sensing system. The basic unit (cluster) consists of four electrochemical cells (EC), each containing one pseudoreference electrode (PRE) and one working electrode (WE). All four ECs are simultaneously interrogated, each at different frequencies, with square wave pulses superposed on a sawtooth signal for cyclic voltammetry (CV). Lock-in amplification provides independent read-out of four signals, with excellent noise suppression. We expanded a single cluster system into an array of 16 clusters by using electronic switches. The chip with an array of ECs was fabricated using planar technology with a gap between a WE and a PRE of ≈2 μm, which results in partial microelectrode-type behavior. The basic electrode characterization was performed with the model case using a ferricyanide–ferrocyanide redox couple (Fe2+/Fe3+) reaction, performing CV and differential pulse voltammetry (DPV). We then used this system to perform cyclic lock-in voltammetry (...

This paper proposes a new method for characterization of 2D materials under the precisely specified conditions. It is achieved by integration of a 2D material as a field effect transistors structures with a piezoelectric resonator. Properties of the 2D material can be mechanically adjusted by the resonator. It results in the independent and precise control of an amplitude of induced mechanical strain, its modulating frequency, which all influence the 2D material properties. The electrical field required to measure 2D material field effect transistors will not be affected by the vibrations, thus giving us a chance to perform the precise measurement of the electrical properties of the 2D material. This approach has a great potential for measuring and monitoring cells, enzymes, nucleic acids, deoxyribonucleic acid and ribonucleic acid. It can be also used for measurement of toxic, combustive or waste gases.

Microfluidic devices typically require complex shapes such as funnels, spirals, splitters, channels with different widths, or customized objects of arbitrary complexity with a smooth transition between these elements. Device layouts are generally designed by software developed for the design of integrated circuits or by general computer-aided design drawing tools. Both methods have their limitations, making these tasks time consuming. Here, a script-based, time-effective method to generate the layout of various microfluidic chips with complex geometries is presented. The present work uses the nanolithography toolbox (NT), a platform-independent software package, which employs parameterized fundamental blocks (cells) to create microscale and nanoscale structures. In order to demonstrate the functionality and efficiency of the NT, a few classical microfluidic devices were designed using the NT and then fabricated in glass/silicon using standard microfabrication techniques and in poly(dimethylsiloxane) using soft lithography as well as more complex techniques used for flow-through calorimetry. In addition, the functionality of a few of the fabricated devices was tested. The powerful method proposed allows the creation of microfluidic devices with complex layouts in an easy way, simplifying the design process and improving design efficiency. Thus, it holds great potential for broad applications in microfluidic device design.

The modification of surface properties frequently requires the binding of suitable compounds to the original surface. Silanes or thiols can be directly covalently bonded to either Si-based materials or Au, thus ruling out polymers. Here, we show the utilization of a layer of SiO2 with a thickness of a few nanometers that serves as a cross-linker between polymers and silanes providing covalent bonding to the surface. We deposited a polymer onto a thermally oxidized microstructured Si surface followed by subsequent Si removal. We demonstrated a Si-based nanotechnology fabrication method that can be generally used to modify the surface properties of practically any polymer via SiO2 cross-linking. This can produce any topology, including microstructures, nanostructures, or composite microstructure/nanostructures terminating in different shapes, since all the steps involving polymer deposition are conducted at room temperature after the Si surface has been thermally oxidized. This technique opens a broad field of new applications for polymers in microstructures and nanostructures that have stable water surface contact angle values with the contact angle set by demand for gecko-mimicking structures or lotus leaf inspired surfaces.

Here we report one of the smallest real-time polymerase chain reaction (PCR) system up to date with approximate size of 100 mm × 60 mm × 33 mm. The system is an autonomous unit requiring an external 12 V power supply. Four simultaneous reactions are performed in form of virtual reaction chambers (VRC) where a 200 nL sample is covered with mineral oil and placed on a glass cover slip. Fast, 40 cycle amplification of an amplicon from the H7N9 gene was used to demonstrate PCR performance. The standard curve slope was (-3.02 ± 0.16) cycles at threshold per decade (mean ± standard deviation) corresponding to an amplification efficiency of (0.91 ± 0.05) per cycle (mean ± standard deviation). The PCR device was capable of detecting a single deoxyribonucleic acid (DNA) copy. These results further suggest that our handheld PCR device may have broad, technologically-relevant applications extending to rapid detection of infectious diseases in small clinics. Zde hlásí jeden z nejmenších real-time polymerázové řetězové reakce (PCR), systému, až do dnešního dne s přibližné velikosti 100 mm × 60 mm × 33 mm. Systém je autonomní jednotka vyžaduje externí 12 V napájecí zdroj. Čtyři simultánní Reakce se provádějí ve formě virtuálních reakčních komor (VRC), ve kterých je vzorek nL 200 na které se vztahuje s minerálním olejem a umístí se na skleněné krycí sklíčko. Fast, zesílení 40 cyklus amplikon, z genu H7N9 byl použit pro demonstraci výkonu PCR. Standardní křivka byla sklon (-3,02 ± 0,16) cyklů při prahu za deset let (průměr ± standardní odchylka), což odpovídá účinnosti amplifikace (0,91 ± 0,05) na jeden cyklus (střední hodnota ± standardní odchylka). PCR zařízení bylo schopné detekovat jeden deoxyribonukleové kyseliny (DNA) kopie. Tyto výsledky dále ukazují, že naše kapesní PCR přístroj může mít široké, technologicky příslušných žádostí procházejících rychlou detekci infekčních nemocí v malých klinikách.

In this work, we demonstrate the simple fabrication process of AlN-based piezoelectric energy harvesters (PEH), which are made of cantilevers consisting of a multilayer ion beam-assisted deposition. The preferentially (001) orientated AlN thin films possess exceptionally high piezoelectric coefficients d33 of (7.33 &plusmn, 0.08) pC∙N&minus, 1. The fabrication of PEH was completed using just three lithography steps, conventional silicon substrate with full control of the cantilever thickness, in addition to the thickness of the proof mass. As the AlN deposition was conducted at a temperature of &asymp, 330 &deg, C, the process can be implemented into standard complementary metal oxide semiconductor (CMOS) technology, as well as the CMOS wafer post-processing. The PEH cantilever deflection and efficiency were characterized using both laser interferometry, and a vibration shaker, respectively. This technology could become a core feature for future CMOS-based energy harvesters.

The global risk of viral disease outbreaks emphasizes the need for rapid, accurate, and sensitive detection techniques to speed up diagnostics allowing early intervention. An emerging field of microfluidics also known as the lab-on-a-chip (LOC) or micro total analysis system includes a wide range of diagnostic devices. This review briefly covers both conventional and microfluidics-based techniques for rapid viral detection. We first describe conventional detection methods such as cell culturing, immunofluorescence or enzyme-linked immunosorbent assay (ELISA), or reverse transcription polymerase chain reaction (RT-PCR). These methods often have limited speed, sensitivity, or specificity and are performed with typically bulky equipment. Here, we discuss some of the LOC technologies that can overcome these demerits, highlighting the latest advances in LOC devices for viral disease diagnosis. We also discuss the fabrication of LOC systems to produce devices for performing either individual steps or virus detection in samples with the sample to answer method. The complete system consists of sample preparation, and ELISA and RT-PCR for viral-antibody and nucleic acid detection, respectively. Finally, we formulate our opinions on these areas for the future development of LOC systems for viral diagnostics., Graphical abstract Image 1, Highlights • Conventional methods for viral detection are time consuming and labor intensive. • Newly emerged microfluidic devices can perform practically any conventional laboratory technique. • Here we concentrated on microfluidics-based viral detection by immunoassays as well as by nucleic acid amplification. • We used a few examples of successfully commercialized rapid devices. • We discuss perspectives for technology of viral detection in near future.

We developed an advanced method for fabricating microfluidic structures comprising channels and inputs/outputs buried within a silicon wafer based on single level lithography. We etched trenches into a silicon substrate, covered these trenches with parylene-C, and selectively opened their bottoms using femtosecond laser photoablation, forming channels and inputs/outputs by isotropic etching of silicon by xenon difluoride vapors. We subsequently sealed the channels with a second parylene-C layer. Unlike in previously published works, this entire process is conducted at ambient temperature to allow for integration with complementary metal oxide semiconductor devices for smart readout electronics. We also demonstrated a method of chip cryo-cleaving with parylene presence that allows for monitoring of the process development. We also created an observation window for in situ visualization inside the opaque silicon substrate by forming a hole in the parylene layer at the silicon backside and with local silicon removal by xenon difluoride vapor etching. We verified the microfluidic chip performance by forming a segmented flow of a fluorescein solution in an oil stream. This proposed technique provides opportunities for forming simple microfluidic systems with buried channels at ambient temperature.

The polymerase chain reaction (PCR) method and its quantitative variant (qPCR) were a landmark discovery for detection and quantification of small amounts of unambiguous deoxyribonucleic acids (DNA) due to its enormous sensitivity and specificity. The current methods for qPCR protocol optimization provide no information of the PCR propagation during the cycles as only single data point is extracted at the end of each thermal cycles, limiting a thorough understanding of reaction details. In this study, we utilized the continuous fluorescence monitoring method to observe progress of the reaction in real time with over 100 data points per each cycle, thus gaining a profound insight into the PCR itself. This provided information about the real-time PCR status, dominating reactions and their completion/incompletion during each cycle as well as their reaction kinetics. We then adjusted the duration of either annealing or elongation steps to ensure their completion within each cycle, resulting in the protocol optimization with complete amplification, enhancing PCR efficiency and taking

Here, we report a single-point detection method for the determination of dynamic surface conditions inside microfluidic channels. The proposed method is based on monitoring fluorescence amplitude as a function of the convolution of a laser beam with segmented flow consisting of two immiscible liquids, one containing fluorescent dye. The fluorescence amplitude is determined by the flow rate and the droplet shape, which is affected by the channel surface properties. We modeled the interaction of a droplet and a laser beam via computer-aided design software, using the laser beam location in relation to the droplet shape as a parameter. The method was applied to fused silica capillaries with both unmodified and modified surfaces, with segmented flow exhibiting water contact angles of ≈ 30° and ≈ 100°, respectively. The method allows discrimination between hydrophillic and hydrophobic surfaces, as well as the quality of the treatment. The results were verified using fluorescence imaging of the droplets via a stroboscopic technique. We also applied this method to the analysis of microfabricated channels with non-circular cross sections. We demonstrated that the technique enables the determination of the hydrophobicity of channel surfaces, a crucial property required for the generation of segmented flow or emulsions for applications such as digital PCR.

Herein, we introduce microfluidic superheating as a new method for peptide fragmentation prior to mass spectrometric analysis. The superheating conditions were found to be stable up to 240 °C for more than 30 min without elevated pressure or boiling of the aqueous sample. As proof of principle, we exposed the peptides ACTH1-10 and OVA257-264 to various superheating conditions, causing different degrees of decomposition. Optimized superheating conditions resulted in the entire peptide ladder sequence of the y-ions, allowing the amino acid sequence to be deduced from a single-stage mass spectrum. Thus, obtaining information in the same quality as from tandem mass spectrometry can be achieved by a single superheating step.

In this paper we report a method of continuous electroextraction of amino acids as model samples using a caseinate–poly(ethylene-glycol) – PEG – two phase system in a microchip able to separate compounds based on their differences in electrophoretic mobility and solvent affinity. Fundamentally, the phase boundary replaces a physical membrane, suppressing the diffusion. When external potential is applied, the molecules selectively cross this barrier. The selectivity of the amino acids extraction is the result of their electrochemical properties and applied voltage. We applied this method to amino acids extraction and the results suggest the possibility of high levels of purification by controlling the electric field strength across the liquid barrier. It is a promising method for complex sample separation as well as for the purification of individual compounds.

We demonstrate the use of open-surface microfluidics to sequence DNA by pyrosequencing at the plain hydrophobically coated surface of a microscope glass cover slip. This method offers significant advantages in terms of instrument size, simplicity, disposability, and functional integration, particularly when combined with the broad and flexible capabilities of open-surface microfluidics. The DNA was incubated on superparamagnetic particles and placed on a hydrophobically coated glass substrate. The particles with bound DNA were moved using magnetic force through microliter-sized droplets covered with mineral oil to prevent water evaporation from the droplets. These droplets served as reaction “stations” performing pyrosequencing as well as washing stations. The resequencing protocol with 34-mer single-stranded DNA (ssDNA) was used to determine the reaction performance. The de novo sequencing was performed with 51-mer and 81-mer ssDNA. The method can be integrated with previously shown sample preparation and PCR into a single sample-to-answer system on a plain glass surface.

Carbon nanotubes (CNTs) have near unity infrared (IR) absorption efficiency, making them extremely attractive in IR imaging devices. Since CNT growth occurs at elevated temperatures, integration of CNTs with IR imaging devices is challenging and has not yet been achieved. Here we show a strategy for implementing CNTs as IR absorbers using differential heating of thermally-isolated microbolometer membranes in a C(2)H(2) environment. During the process, CNTs were catalytically grown on the surface of a locally-heated membrane while the substrate was maintained at an ambient temperature. CNT growth was monitored in situ in real time using optical microscopy. During growth, we measured the intensity of light emission and the reflected light from the heated microbolometer. Our measurements of bolometer performance show that the CNT layer on the surface of the microbolometer membrane increases the IR response by a factor of (2.3 ± 0.1) (mean ± one standard deviation of the least-squares fit parameters). This work opens the door to integrating near unity IR absorption, CNT-based, IR absorbers with hybrid complementary metal-oxide-semiconductor focal plane array architectures.

The invention of polymerase chain reaction (PCR) in 1983 revolutionized many areas of science, due to its ability to multiply a number of copies of DNA sequences (known as amplicons). Here we report on a method to double the throughput of quantitative PCR which could be especially useful for PCR-based mass screening. We concurrently amplified two target genes using only single fluorescent dye. A FAM probe labelled olionucleotide was attached to a quencher for one amplicon while the second one was without a probe. The PCR was performed in the presence of the intercalating dye SYBR Green I. We collected the fluorescence amplitude at two points per PCR cycle, at the denaturation and extension steps. The signal at denaturation is related only to the amplicon with the FAM probe while the amplitude at the extension contained information from both amplicons. We thus detected two genes within the same well using a single fluorescent channel. Any commercial real-time PCR systems can use this method doubling the number of detected genes. The method can be used for absolute quantification of DNA using a known concentration of housekeeping gene at one fluorescent channel.

We report on a microfluidic system formed by 200 nl water droplets, encapsulated by a 600 nl mineral oil placed on a hydrophobically coated glass microscope cover slip. The micromachined heater underneath the glass was able to heat up the sample at a heating rate of 650 C/s, heating the water sample up to 200 C in less than 2 s. The sample/glass and the sample/oil interface did not have nucleation centers, showing that the sample reached a superheated stage without the necessity of being pressurized to suppress boiling. This method can be utilized for various applications currently being conducted in autoclaves. Popisujeme mikrofluidní systém tvořeném 200 nl kapou vody, pokryte 600 nl minerálniho oleje umístěn na hydrofobně potažené skleněné mikroskopické krycí sklíčko.micromachined topné těleso pod sklem byl schopen ohřát vzorku při rychlosti zahřívání 650? C / s, topení Vzorek vody až do 200? C, za méně než 2 s.Vzorek / skla a vzorek / olej rozhraní ne mají nukleační centra, což ukazuje, že vzorek dosáhl přehřáté fáze bez nutnosti být pod tlakem potlačit varu. Tato metoda může být použita pro různé aplikace v současné době se provádí v autoklávech.

We present a portable, battery-operated and application-specific lab-on-a-chip (ASLOC) system that can be easily configured for a wide range of lab-on-a-chip applications. It is based on multiplexed electrical current detection that serves as the sensing system. We demonstrate different configurations to perform most detection schemes currently in use in LOC systems, including some of the most advanced such as nanowire-based biosensing, surface plasmon resonance sensing, electrochemical detection and realtime PCR. The complete system is controlled by a single chip and the collected information is stored in situ, with the option of transferring the data to an external display by using a USB interface. In addition to providing a framework for truly portable real-life developments of LOC systems, we envisage that this system will have a significant impact on education, especially since it can easily demonstrate the benefits of integrated microanalytical systems. Představujeme přenosný, Lab-on-a-chip (ASLOC) bateriový systém ovládaná a pro konkrétní aplikaci, které mohou snadno nakonfigurovat pro široký rozsah lab-on-a-chip aplikací. Je založen na multiplexovaný elektrické detekce proudu, která slouží jako snímací systém. Demonstrujeme různé konfigurace provádět Většina systémů detekce v současné době použití v LOC systémech, včetně těch nejvyspělejších, jako je nanowire bázi biosnímací, povrch plasmon rezonance snímání, elektrochemické detekce a realtime PCR. Celý systém je řízen jediném čipu a Shromážděné informace je uložena in situ, s možností přenosu dat na externí displej pomocí rozhraní USB. Kromě toho se poskytne rámec pro skutečně přenosných reálného života vývoji LOC systémů, můžeme předpokládat, že tento Systém bude mít významný dopad na vzdělávání, zejména proto, že lze snadno prokázat výhody integrovaných mikroanalytické systémů.