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Amino acids (AAs) have broad nutritional, therapeutic, and medical significance and thus are one of the most common active ingredients of nutritional supplements. Analytical strategies for determining AAs are high-priced and often limited to methods that require modification of AA polarity or incorporation of an aromatic moiety. The aim of this work was to develop a new method for the determination of L-arginine, L-ornithine, and L-lysine on low-cost microchip electrophoresis instrumentation conjugated with capacitively coupled contactless conductivity detection. A solution consisting of 0.3 M acetic acid and 1 × 10−5 M iminodiacetic acid has been identified as the optimal background electrolyte, ensuring the shortest possible analysis time. The short migration times of amino acids (t ≤ 64 s) and method simplicity resulted in high analysis throughput with high precision and linearity (R2 ≥ 0.9971). The limit of detection values ranged from 0.15 to 0.19 × 10−6 M. The accuracy of the proposed method was confirmed by recovery measurements. The results were compared with CE-UV-VIS and HPLC-DAD methods and showed good agreement. This work represents the first successful demonstration of the ME-C4D analysis of L-arginine, L-ornithine, and L-lysine in real samples. [ABSTRACT FROM AUTHOR]

The growing consumption of plant-based milk substitutes raises important questions about their composition. The various additives used by manufacturers, including those employed as flavor enhancers, protein additives, and stabilizers, may contain both protein and non-protein nitrogen components. In our study, we examined not only popular milk alternatives but also other milk substitutes made from specific plants. We present a reproducible and rapid method for the simultaneous qualitative and quantitative determination of the total nitrogen content in milk alternatives, focusing on applicability. Using the microchip gel electrophoretic method, we determined that the total nitrogen content differed from the protein content indicated on the packaging. Our results, along with statistical evaluations, supported the hypothesis that different brands of products, derived from the same plant source, resulted in different microfluidic profiles, likely due to the presence of additives. As expected, the microfluidic profiles of additive-free products differed from those of fortified products made from the same plant-based milk replacer. Total nitrogen content provides crucial information for individuals with kidney disease, as is essential to reduce the burden on the kidneys to slow deterioration, alleviate symptoms and avoid complications. [ABSTRACT FROM AUTHOR]

Hemoglobin (Hb) disorders are among the most common monogenic diseases affecting nearly 7% of the world population. Among various Hb disorders, approximately 1.5% of the world population carries β-thalassemia (β-Thal), affecting 40,000 newborns every year. Early screening and a timely diagnosis are essential for β-thalassemia patients for the prevention and management of later clinical complications. However, in Africa, Southern Europe, the Middle East, and Southeast Asia, where β-thalassemia is most prevalent, the diagnosis and screening for β-thalassemia are still challenging due to the cost and logistical burden of laboratory diagnostic tests. Here, we present Gazelle, which is a paper-based microchip electrophoresis platform that enables the first point-of-care diagnostic test for β-thalassemia. We evaluated the accuracy of Gazelle for the β-Thal screening across 372 subjects in the age range of 4–63 years at Apple Diagnostics lab in Mumbai, India. Additionally, 30 blood samples were prepared to mimic β-Thal intermediate and β-Thal major samples. Gazelle-detected levels of Hb A, Hb F, and Hb A2 demonstrated high levels of correlation with the results reported through laboratory gold standard high-performance liquid chromatography (HPLC), yielding a Pearson correlation coefficient = 0.99. This ability to obtain rapid and accurate results suggests that Gazelle may be suitable for the large-scale screening and diagnosis of β-Thal. [ABSTRACT FROM AUTHOR]

The health supplement industry is one of the fastest growing industries in the world, but there is a lack of suitable analytical methods for the determination of active compounds in health supplements such as peptides. The present work describes an implementation of contactless conductivity detection on microchip technology as a new strategy for the electrophoretic determination of L-carnosine in complex health supplement formulations without pre-concentration and derivatization steps. The best results were obtained in the case of +1.00 kV applied for 20 s for injection and +2.75 kV applied for 260 s for the separation step. Under the selected conditions, a linear detector response of 5 × 10−6 to 5 × 10−5 M was achieved. L-carnosine retention time was 61 s. The excellent reproducibility of both migration time and detector response confirmed the high precision of the method. The applicability of the method was demonstrated by the determination of L-carnosine in three different samples of health supplements. The recoveries ranged from 91 to 105%. Subsequent analysis of the samples by CE-UV-VIS and HPLC-DAD confirmed the accuracy of the obtained results. [ABSTRACT FROM AUTHOR]

The problem of overlapping peaks has been a challenge in microchip electrophoresis (ME) signal analysis. However, traditional peak fitting algorithms have difficulty analyzing overlapping peaks due to the high dependence on the starting point. In this study, we propose a symmetrical peak fitting method named the tent-mapped whale optimization algorithm and Levenberg–Marquardt (TWOALM), which combines a whale optimization algorithm (WOA) improved by one of the most commonly used chaotic maps, the tent map and the Levenberg–Marquardt (LM) algorithm. Specifically, we first derive the fitted model for the overlapping peaks, showing that it is separable nonlinear least squares, significantly reducing the number of parameters to be optimized. Second, we integrate the tent map into the WOA, which improves the convergence speed of the peak fitting algorithm. Finally, we propose an efficient peak-fitting algorithm that combines the improved WOA and LM. The advantage of the proposed algorithm is that it is significantly faster than WOA and significantly more accurate than the LM algorithm. The results of fitting the synthetic peaks and ME signals showed that the combination of the chaotic map-based WOA algorithm and the LM algorithm can significantly improve the peak fitting performance and provide an effective solution for the analysis of overlapping peaks. [ABSTRACT FROM AUTHOR]

Nitrite is considered an important target analyte for environmental monitoring. In water resources, nitrite is the result of the nitrogen cycle and the leaching processes of pesticides based on nitrogenous compounds. A high concentration of nitrite can be associated with intoxication processes and metabolic disorders in humans. The present study describes the development of a portable analytical methodology based on microchip electrophoresis coupled with amperometric detection for the determination of nitrite in environmental water samples. Electrophoretic and detection conditions were optimized, and the best separations were achieved within 60 s by employing a mixture of 30 mmol L−1 lactic acid and 15 mmol L−1 histidine (pH = 3.8) as a running buffer applying 0.7 V to the working electrode (versus Pt) for amperometric measurements. The developed methodology revealed a satisfactory linear behavior in the concentration range between 20 and 80 μmolL−1 (R2 = 0.999) with a limit of detection of 1.3 μmolL−1. The nitrite concentration was determined in five water samples and the achieved values ranged from (28.7 ± 1.6) to (67.1 ± 0.5) µmol L−1. The data showed that using the proposed methodology revealed satisfactory recovery values (83.5–103.8%) and is in good agreement with the reference technique. Due to its low sample consumption, portability potential, high analytical frequency, and instrumental simplicity, the developed methodology may be considered a promising strategy to monitor and quantitatively determine nitrite in environmental samples. [ABSTRACT FROM AUTHOR]

Initial screening of criminal evidence often involves serological testing of stains of unknown composition and/or origin discovered at a crime scene to determine the tissue of origin. This testing is presumptive but critical for contextualizing the scene. Here, we describe a microfluidic approach for body fluid profiling via fluorescent electrophoretic separation of a published mRNA panel that provides unparalleled specificity and sensitivity. This centrifugal microfluidic approach expedites and automates the electrophoresis process by allowing for simple, rotationally driven flow and polymer loading through a 5 cm separation channel; with each disc containing three identical domains, multi-sample analysis is possible with a single disc and multi-sample detection per disc. The centrifugal platform enables a series of sequential unit operations (metering, mixing, aliquoting, heating, storage) to execute automated electrophoretic separation. Results show on-disc fluorescent detection and sizing of amplicons to perform comparably with a commercial 'gold standard' benchtop instrument and permitted sensitive, empirical discrimination between five distinct body fluids in less than 10 min. Notably, our microfluidic platform represents a faster, simpler method for separation of a transcriptomic panel to be used for forensically relevant body fluid identification. [ABSTRACT FROM AUTHOR]

Keywords: lab-on-PCB; printed circuit board; biomedical applications; electronics; sensors; biosensors; actuators; lab-on-chip; microfluidics EN lab-on-PCB printed circuit board biomedical applications electronics sensors biosensors actuators lab-on-chip microfluidics N.PAG N.PAG 4 08/03/22 20220701 NES 220701 The use of Printed Circuit Boards (PCBs) has seen a remarkable growth over the last decade, with applications in engineering, medicine, biology, chemistry, etc. The lab-on-PCB devices are an interesting improvement of lab-on-chip devices due to the inexpensive characteristic of Printed Circuit Boards, and the possibility of including microfluidic circuits, sensors, and actuators, together with electronics in the same substrate. Therefore, the achievement of a robust development of biosensor using PCB substrates is mandatory for a successful future of lab-on-PCB devices. Apart from sensors and biosensors, the lab-on-PCB devices can include actuators, such as microheaters [[4]], and devices for microfluid handling, for example, electrowetting on dielectric devices (EWOD) [[5]]. [Extracted from the article]

In the microchip electrophoresis with capacitively coupled contactless conductivity detection, the stray capacitance of the detector causes high background noise, which seriously affects the sensitivity and stability of the detection system. To reduce the effect, a novel design of planar grounded capacitively coupled contactless conductivity detector (PG-C4D) based on printed circuit board (PCB) is proposed. The entire circuit plane except the sensing electrodes is covered by the ground electrode, greatly reducing the stray capacitance. The efficacy of the design has been verified by the electrical field simulation and the electrophoresis detection experiments of inorganic ions. The baseline intensity of the PG-C4D was less than 1/6 of that of the traditional C4D. The PG-C4D with the new design also demonstrated a good repeatability of migration time, peak area, and peak height (n = 5, relative standard deviation, RSD ≤ 0.3%, 3%, and 4%, respectively), and good linear coefficients within the range of 0.05–0.75 mM (R2 ≥ 0.986). The detection sensitivity of K+, Na+, and Li+ reached 0.05, 0.1, and 0.1 mM respectively. Those results prove that the new design is an effective and economical approach which can improve sensitivity and repeatability of a PCB based PG-C4D, which indicate a great application potential in agricultural and environmental monitoring. [ABSTRACT FROM AUTHOR]

Cell culture and cell scaffold engineering have previously developed in two directions. First can be 'static into dynamic', with proven effects that dynamic cultures have benefits over static ones. Researches in this direction have used several mechanical means, like external vibrators or shakers, to approximate the dynamic environments in real tissue, though such approaches could only partly address the issue. Second, can be '2D into 3D', that is, artificially created three-dimensional (3D) passive (also called 'static') scaffolds have been utilized for 3D cell culture, helping external culturing conditions mimic real tissue 3D environments in a better way as compared with traditional two-dimensional (2D) culturing. In terms of the fabrication of 3D scaffolds, 3D printing (3DP) has witnessed its high popularity in recent years with ascending applicability, and this tendency might continue to grow along with the rapid development in scaffold engineering. In this review, we first introduce cell culturing, then focus 3D cell culture scaffold, vibration stimulation for dynamic culture, and 3DP technologies fabricating 3D scaffold. Potential interconnection of these realms will be analyzed, as well as the limitations of current 3D scaffold and vibration mechanisms. In the recommendation part, further discussion on future scaffold engineering regarding 3D vibratory scaffold will be addressed, indicating 3DP as a positive bridging technology for future scaffold with integrated and localized vibratory functions. [ABSTRACT FROM AUTHOR]

We investigated an anomalous DNA separation where two DNA fragments from the human Y-chromosome sY638 (64 bp) and sY592 (65 bp), with only one base pair difference, were separated. This result is abnormal since in a previous study, we found that 5 bp was the minimum difference between two DNA fragments that the microchip electrophoresis system can separate. The formation of a mini-loop in the structure of the DNA fragment of sY638 (64 bp) was strongly expected to be the reason. To investigate this, we synthesized three modified DNA fragments for sY638 (64 bp), and the modifications were in two expected locations for possible mini-loop formation. Later, the separation between sY592 (65 bp) and the three modified fragments of sY638 (64 bp) was not possible. Thus, we conclude that the formation of a mini-loop in the structure of the DNA is the reason behind this anomalous separation. [ABSTRACT FROM AUTHOR]

A micro surface tension pump is a new type of low-cost, built-in, all-glass, microfluidic pump on a glass microchip fabricated by one-step glass etching. However, geometric minimization and optimization for practical use are challenging. Here, we report a one-square-millimeter, built-in, all-glass pump controlled by two-way digital gas pressure. The pump consists simply of two joint chambers and a piston between two gas control channels. It does not require pre-perfusion for initialization, and can immediately begin to run when a liquid enters its inlet channel. It is also more reliable than conventional micro pumps for practical use due to its ability to restart after the formation of a blocking bubble, which can serve as a valuable troubleshooting procedure. Its volumetric pump output was 0.5-0.7 nL· s-1 under a pump head pressure of 300 Pa. [ABSTRACT FROM AUTHOR]

Soft micro devices and stretchable electronics have attracted great interest for their potential applications in sensory skins and wearable bio-integrated devices. One of the most important steps in building printed circuits is the alignment of assembled micro objects. Previously, the capillary self-alignment of microchips driven by surface tension effects has been shown to be able to achieve high-throughput and high-precision in the integration of micro parts on rigid hydrophilic/superhydrophobic patterned surfaces. In this paper, the self-alignment of microchips on a patterned soft and stretchable substrate, which consists of hydrophilic pads surrounded by a superhydrophobic polydimethylsiloxane (PDMS) background, is demonstrated for the first time. A simple process has been developed for making superhydrophobic soft surface by replicating nanostructures of black silicon onto a PDMS surface. Different kinds of PDMS have been investigated, and the parameters for fabricating superhydrophobic PDMS have been optimized. A self-alignment strategy has been proposed that can result in reliable self-alignment on a soft PDMS substrate. Our results show that capillary self-alignment has great potential for building soft printed circuits. [ABSTRACT FROM AUTHOR]

We propose a system that transports oocytes and measures their mechanical characteristics in an open environment using a robot integrated microfluidic chip (chip). The cells are transported through a micropillar array in the chip, and their characteristics are measured by a mechanical probe and a force sensor. Because the chip has an open microchannel, important cells such as oocytes are easily introduced and collected without the risk for losing them. In addition, any bubbles trapped in the chip, which degrade the measurement precision, are easily removed. To transport the oocytes through the open microchannel, we adopt a transportation technique based on a vibration-induced flow. Under this flow, oocytes arrive at the measurement point, where their mechanical characteristics are determined. We demonstrate the introduction, transportation, measurement of mechanical characteristics, and collection of oocytes using this system. [ABSTRACT FROM AUTHOR]

Single-cell analysis has become the interest of a wide range of biological and biomedical engineering research. It could provide precise information on individual cells, leading to important knowledge regarding human diseases. To perform single-cell analysis, it is crucial to isolate the individual cells before further manipulation is carried out. Recently, microfluidic biochips have been widely used for cell trapping and single cell analysis, such as mechanical and electrical detection. This work focuses on developing a finite element simulation model of single-cell trapping system for any types of cells or particles based on the hydrodynamic flow resistance (Rh) manipulations in the main channel and trap channel to achieve successful trapping. Analysis is carried out using finite element ABAQUS-FEAΓ software. A guideline to design and optimize single-cell trapping model is proposed and the example of a thorough optimization analysis is carried out using a yeast cell model. The results show the finite element model is able to trap a single cell inside the fluidic environment. Fluid's velocity profile and streamline plots for successful and unsuccessful single yeast cell trapping are presented according to the hydrodynamic concept. The single-cell trapping model can be a significant important guideline in designing a new chip for biomedical applications. [ABSTRACT FROM AUTHOR]

Advances in biotechnology have increased the demand for suitable analytical techniques for the analysis of genetically modified organisms. Study of the substantial equivalence, discrimination between transgenic and non-transgenic cultivars, study of the unintended effects caused by a genetic modification or their response to diverse situations or stress conditions (e.g., environmental, climatic, infections) are some of the concerns that need to be addressed. Capillary electrophoresis (CE) is emerging as an alternative to conventional techniques for the study and characterization of genetically modified organisms. This article reviews the most recent applications of CE for the analysis and characterization of transgenic cultivars in the last five years. Different strategies have been described depending on the level analyzed (DNA, proteins or metabolites). Capillary gel electrophoresis (CGE) has shown to be particularly useful for the analysis of DNA fragments amplified by PCR. Metabolites and proteins have been mainly separated using capillary zone electrophoresis (CZE) using UV and MS detection. Electrophoretic chips have also proven their ability in the analysis of transgenic cultivars and a section describing the new applications is also included. [ABSTRACT FROM AUTHOR]

The analysis of a microRNA (miRNA), miR-222 isolated from the PC12 cell line, was performed by use of the ribonuclease (RNase) protection assay, cyanine 5 (Cy5)-labeled miR-222 riboprobe, and a Hitachi SV1210 microchip electrophoresis system, which can be used to evaluate the integrity of total RNA. The fluorescence intensity corresponding to the protected RNA fragment increased in a dose-dependent manner with respect to the complementary-strand RNA. More highly sensitive detection of miRNA by microchip electrophoresis than by conventional method using fluorescence-labeled riboprobe could be obtained in 180 s. An obvious increase in miR-222 expression induced by nerve growth factor in PC12 cells could be observed. These results clearly indicate the potential of microchip electrophoresis for the analysis of miRNA using RNase protection assay with a fluorescence-labeled riboprobe. [ABSTRACT FROM AUTHOR]

Paper-based microchip electrophoresis has the potential to bring laboratory electrophoresis tests to the point of need. However, high electric potential and current values induce pH and temperature shifts, which may affect biomolecule electrophoretic mobility thus decrease test reproducibility and accuracy of paper-based microfluidic electrophoresis. We have previously developed a microchip electrophoresis system, HemeChip, which has the capability of providing low-cost, rapid, reproducible, and accurate point-of-care (POC) electrophoresis tests for hemoglobin analysis. Here, we report the methodologies we implemented for characterizing HemeChip system pH and temperature during the development process, including utilizing commercially available universal pH indicator and digital camera pH shift characterization, and infrared camera characterizing temperature shift characterization. The characterization results demonstrated that pH shifts up to 1.1 units, a pH gradient up to 0.11 units/mm, temperature shifts up to 40 °C, and a temperature gradient up to 0.5 °C/mm existed in the system. Finally, we report an acid pre-treatment of the separation media, a cellulose acetate paper, mitigated both pH and temperature shifts and provided a stable environment for reproducible HemeChip hemoglobin electrophoresis separation. [ABSTRACT FROM AUTHOR]

Microfluidics is the advanced microtechnology of fluid manipulation in channels with at least one dimension in the range of 1–100 microns. Microfluidic technology offers a growing number of tools for manipulating small volumes of fluid to control chemical, biological, and physical processes relevant to separation, analysis, and detection. Currently, microfluidic devices play an important role in many biological, chemical, physical, biotechnological and engineering applications. There are numerous ways to fabricate the necessary microchannels and integrate them into microfluidic platforms. In peptidomics and proteomics, microfluidics is often used in combination with mass spectrometric (MS) analysis. This review provides an overview of using microfluidic systems for peptidomics, proteomics and cell analysis. The application of microfluidics in combination with MS detection and other novel techniques to answer clinical questions is also discussed in the context of disease diagnosis and therapy. Recent developments and applications of capillary and microchip (electro)separation methods in proteomic and peptidomic analysis are summarized. The state of the art of microchip platforms for cell sorting and single-cell analysis is also discussed. Advances in detection methods are reported, and new applications in proteomics and peptidomics, quality control of peptide and protein pharmaceuticals, analysis of proteins and peptides in biomatrices and determination of their physicochemical parameters are highlighted. [ABSTRACT FROM AUTHOR]

Carnosine (β-alanyl-L-histidine) is a naturally occurring endogenous peptide widely distributed in excitable tissues such as the brain. This dipeptide has well-known antioxidant, anti-inflammatory, and anti-aggregation activities, and it may be useful for treatment of neurodegenerative disorders such as Alzheimer's disease (AD). In this disease, peripheral infiltrating macrophages play a substantial role in the clearance of amyloid beta (Aβ) peptides from the brain. Correspondingly, in patients suffering from AD, defects in the capacity of peripheral macrophages to engulf Aβ have been reported. The effects of carnosine on macrophages and oxidative stress associated with AD are consequently of substantial interest for drug discovery in this field. In the present work, a model of stress induced by Aβ1-42 oligomers was investigated using a combination of methods including trypan blue exclusion, microchip electrophoresis with laser-induced fluorescence, flow cytometry, fluorescence microscopy, and high-throughput quantitative real-time PCR. These assays were used to assess the ability of carnosine to protect macrophage cells, modulate oxidative stress, and profile the expression of genes related to inflammation and pro- and antioxidant systems. We found that pre-treatment of RAW 264.7 macrophages with carnosine counteracted cell death and apoptosis induced by Aβ1-42 oligomers by decreasing oxidative stress as measured by levels of intracellular nitric oxide (NO)/reactive oxygen species (ROS) and production of peroxynitrite. This protective activity of carnosine was not mediated by modulation of the canonical inflammatory pathway but instead can be explained by the well-known antioxidant and free-radical scavenging activities of carnosine, enhanced macrophage phagocytic activity, and the rescue of fractalkine receptor CX3CR1. These new findings obtained with macrophages challenged with Aβ1-42 oligomers, along with the well-known multimodal mechanism of action of carnosine in vitro and in vivo, substantiate the therapeutic potential of this dipeptide in the context of AD pathology. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus (S. aureus) is one of the most common pathogens for nosocomial and community infections, which is closely related to the occurrence of pyogenic and toxic diseases in human beings. In the current study, a lab-built microchip capillary electrophoresis (microchip CE) system was employed for the rapid determination of S. aureus, while a simple-to-use space domain internal standard (SDIS) method was carried out for the reliable quantitative analysis. The precision, accuracy, and reliability of SDIS were investigated in detail. Noted that these properties could be elevated in SDIS compared with traditional IS method. Remarkably, the PCR products of S. aureusnuc gene could be identified and quantitated within 80 s. The theoretical detection limit could achieve a value of 0.066 ng/μL, determined by the using SDIS method. The current work may provide a promising detection strategy for the high-speed and highly efficient analysis of pathogens in the fields of food safety and clinical diagnosis. [ABSTRACT FROM AUTHOR]

The last decades of biological, toxicological, and pharmacological research have deeply changed the way researchers select the most appropriate 'pre-clinical model'. The absence of relevant animal models for many human diseases, as well as the inaccurate prognosis coming from 'conventional' pre-clinical models, are among the major reasons of the failures observed in clinical trials. This evidence has pushed several research groups to move more often from a classic cellular or animal modeling approach to an alternative and broader vision that includes the involvement of microfluidic-based technologies. The use of microfluidic devices offers several benefits including fast analysis times, high sensitivity and reproducibility, the ability to quantitate multiple chemical species, and the simulation of cellular response mimicking the closest human in vivo milieu. Therefore, they represent a useful way to study drug–organ interactions and related safety and toxicity, and to model organ development and various pathologies 'in a dish'. The present review will address the applicability of microfluidic-based technologies in different systems (2D and 3D). We will focus our attention on applications of microchip electrophoresis (ME) to biological and toxicological studies as well as in drug discovery and development processes. These include high-throughput single-cell gene expression profiling, simultaneous determination of antioxidants and reactive oxygen and nitrogen species, DNA analysis, and sensitive determination of neurotransmitters in biological fluids. We will discuss new data obtained by ME coupled to laser-induced fluorescence (ME-LIF) and electrochemical detection (ME-EC) regarding the production and degradation of nitric oxide, a fundamental signaling molecule regulating virtually every critical cellular function. Finally, the integration of microfluidics with recent innovative technologies—such as organoids, organ-on-chip, and 3D printing—for the design of new in vitro experimental devices will be presented with a specific attention to drug development applications. This 'composite' review highlights the potential impact of 2D and 3D microfluidic systems as a fast, inexpensive, and highly sensitive tool for high-throughput drug screening and preclinical toxicological studies. [ABSTRACT FROM AUTHOR]