Single-Cell Analysis of Highly Metastatic Circulating Tumor Cells by Combining a Self-Folding Induced Release Reaction with a Cell Capture Microchip.

Nondestructive analysis of the single-cell molecular phenotype of circulating tumor cells (CTCs) is of great significance to the precise diagnosis and treatment of cancer but is also a huge challenge. To address this issue, here, we develop a facile analysis system that integrates CTCs' capture and molecular phenotype analysis. An isothermal nucleic acid amplification technique named self-folding induced release reaction (sFiR), which has high-efficiency signal amplification capabilities and can run under physiological conditions, is first developed to meet the high requirements for sensitivity and nondestructivity. By combining the sFiR with immune recognition and a single cell capture microchip, the molecular phenotype analysis of a single CTC is realized. As a model, nondestructive analysis of junction plakoglobin (JUP), an overexpressed membrane protein that is closely related to the metastasis of CTCs, is successfully achieved. Results reveal that this sFiR-based analysis system can clearly distinguish the expression of JUP in different cancer cell lines and can present quantitative information on the expression of JUP. Furthermore, the captured and analyzed CTCs maintain their basic physiological activity and can be used for drug sensitivity testing. Considering the excellent performance and ease of operation of the system, it can provide technical support for CTC-based cancer liquid biopsy and drug development.

Osteosarcoma: a review of current and future therapeutic approaches.

Osteosarcoma (OS) is the most common primary bone malignancy that affects children and young adults. OS is characterized by a high degree of malignancy, strong invasiveness, rapid disease progression, and extremely high mortality rate; it is considered as a serious threat to the human health globally. The incidence of OS is common in the metaphysis of long tubular bones, but rare in the spine, pelvis, and sacrum areas; moreover, majority of the OS patients present with only a single lesion. OS has a bimodal distribution pattern, that is, its incidence peaks in the second decade of life and in late adulthood. We examine historical and current literature to present a succinct review of OS. In this review, we have discussed the types, clinical diagnosis, and modern and future treatment methods of OS. The purpose of this article is to inspire new ideas to develop more effective therapeutic options.

Organ-on-a-chip: recent breakthroughs and future prospects.

The organ-on-a-chip (OOAC) is in the list of top 10 emerging technologies and refers to a physiological organ biomimetic system built on a microfluidic chip. Through a combination of cell biology, engineering, and biomaterial technology, the microenvironment of the chip simulates that of the organ in terms of tissue interfaces and mechanical stimulation. This reflects the structural and functional characteristics of human tissue and can predict response to an array of stimuli including drug responses and environmental effects. OOAC has broad applications in precision medicine and biological defense strategies. Here, we introduce the concepts of OOAC and review its application to the construction of physiological models, drug development, and toxicology from the perspective of different organs. We further discuss existing challenges and provide future perspectives for its application.

A fully portable microchip real-time polymerase chain reaction for rapid detection of pathogen.

Point-of-care detection for pathogen is of critical need for wide epidemic warning and medical diagnosis. In this work, we have designed and developed a fully portable and integrated microchip based real-time polymerase chain reaction machine for rapid pathogen detection. The instrument consists of three functional components including heating, optical, and electrical modules, which are integrated into a portable compact box. The microchip is consumable material replaceable to meet various detection needs. Consequently, we demonstrated the outstanding performance of this portable machine for rapid detection of Salmonella and Escherichia coli O157:H7 with the advantage of time-saving (∼25 min), less samples consumption, portability, and user-friendly operation.

The Effect of Additives on the Early Stages of Growth of Calcite Single Crystals.

As crystallization processes are often rapid, it can be difficult to monitor their growth mechanisms. In this study, we made use of the fact that crystallization proceeds more slowly in small volumes than in bulk solution to investigate the effects of the soluble additives Mg2+ and poly(styrene sulfonate) (PSS) on the early stages of growth of calcite crystals. Using a "Crystal Hotel" microfluidic device to provide well-defined, nanoliter volumes, we observed that calcite crystals form via an amorphous precursor phase. Surprisingly, the first calcite crystals formed are perfect rhombohedra, and the soluble additives have no influence on the morphology until the crystals reach sizes of 0.1-0.5 µm for Mg2+ and 1-2 µm for PSS. The crystals then continue to grow to develop morphologies characteristic of these additives. These results can be rationalized by considering additive binding to kink sites, which is consistent with crystal growth by a classical mechanism.

3D Droplet Microfluidic Systems for High-Throughput Biological Experimentation.

Herein, we describe the development of a multilayer droplet microfluidic system for creating concentration gradients and generating microdroplets of varying composition for high-throughput biochemical and cell-based screening applications. The 3D droplet-based microfluidic device consists of multiple PDMS layers, which are used to generate logarithmic concentration gradient reagent profiles. Parallel flow focusing structures are used to form picoliter-sized droplets of defined volumes but of varying composition. As proof of concept, we demonstrate rapid enzymatic activity assays and drug cytotoxicity assays on bacteria. The 3D droplet-based microfluidic platform has the potential to allow for high-efficiency and high-throughput analysis, overcoming the structural limitations of single layer microfluidic systems.

Label-free in-flow detection of single DNA molecules using glass nanopipettes.

With the view of enhancing the functionality of label-free single molecule nanopore-based detection, we have designed and developed a highly robust, mechanically stable, integrated nanopipette-microfluidic device which combines the recognized advantages of microfluidic systems and the unique properties/advantages of nanopipettes. Unlike more typical planar solid-state nanopores, which have inherent geometrical constraints, nanopipettes can be easily positioned at any point within a microfluidic channel. This is highly advantageous, especially when taking into account fluid flow properties. We show that we are able to detect and discriminate between DNA molecules of varying lengths when motivated through a microfluidic channel, upon the application of appropriate voltage bias across the nanopipette. The effects of applied voltage and volumetric flow rates have been studied to ascertain translocation event frequency and capture rate. Additionally, by exploiting the advantages associated with microfluidic systems (such as flow control and concomitant control over analyte concentration/presence), we show that the technology offers a new opportunity for single molecule detection and recognition in microfluidic devices.

Electrorheological fluid and its applications in microfluidics.

Microfluidics is a low-cost technique for fast-diagnosis and microsynthesis. Within a decade it might become the foundation of point-of-care and lab-on-a-chip applications. With microfluidic chips, high-throughput sample screening and information processing are made possible. The picoliter droplet runs in microfluidic chips are ideal miniaturized vessels for microdetection and microsynthesis. Meanwhile, individual manipulation of microdroplets remains a challenge: the shortcomings in automatic, reliable, and scalable methods for logic control prevent further integration of microfluidic applications. The giant electrorheological fluid (GERF), which is a kind of "smart" colloid, has tunable viscosity under the influence of external electric field. Therefore, GERF is introduced as the active controlling medium, with real-time response in on-chip fluid control. This review article introduces the working principles and fabrication methods of different types of electrorheological fluid, and extensively describes the strategies of GERF-assisted microfluidic controlling schemes.

An introduction to micro-ecology patches.

Bacterial systems offer excellent tests of how well the general theoretical predictions of ecology dynamics do or do not in fact conform to reality. We believe that the basic rules that govern the cohabitation of competing species for limited resources are the same from bacteria to man, we just don't know the rules, and that fundamental studies of the games bacteria play will give fundamental insight into the vastly more complex systems we hope to attack later. In this tutorial review we discuss how simplified micro-ecologies constructed using tools of micro and nanofabrication techniques offer some idea of how physical principles and analysis can address the issue of complex ecology dynamics.

Investigating the Regulation of Neural Differentiation and Injury in PC12 Cells Using Microstructure Topographic Cues.

In this study, we designed and manufactured a series of different microstructure topographical cues for inducing neuronal differentiation of cells in vitro, with different topography, sizes, and structural complexities. We cultured PC12 cells in these microstructure cues and then induced neural differentiation using nerve growth factor (NGF). The pheochromocytoma cell line PC12 is a validated neuronal cell model that is widely used to study neuronal differentiation. Relevant markers of neural differentiation and cytoskeletal F-actin were characterized. Cellular immunofluorescence detection and axon length analysis showed that the differentiation of PC12 cells was significantly different under different isotropic and anisotropic topographic cues. The expression differences of the growth cone marker growth-associated protein 43 (GAP-43) and sympathetic nerve marker tyrosine hydroxylase (TH) genes were also studied in different topographic cues. Our results revealed that the physical environment has an important influence on the differentiation of neuronal cells, and 3D constraints could be used to guide axon extension. In addition, the neurotoxin 6-hydroxydopamine (6-OHDA) was used to detect the differentiation and injury of PC12 cells under different topographic cues. Finally, we discussed the feasibility of combining the topographic cues and the microfluidic chip for neural differentiation research.

Point-of-care testing detection methods for COVID-19.

COVID-19 is an acute respiratory disease caused by SARS-CoV-2, which has high transmissibility. People infected with SARS-CoV-2 can develop symptoms including cough, fever, pneumonia and other complications, which in severe cases could lead to death. In addition, a proportion of people infected with SARS-CoV-2 may be asymptomatic. At present, the primary diagnostic method for COVID-19 is reverse transcription-polymerase chain reaction (RT-PCR), which tests patient samples including nasopharyngeal swabs, sputum and other lower respiratory tract secretions. Other detection methods, e.g., isothermal nucleic acid amplification, CRISPR, immunochromatography, enzyme-linked immunosorbent assay (ELISA) and electrochemical sensors are also in use. As the current testing methods are mostly performed at central hospitals and third-party testing centres, the testing systems used mostly employ large, high-throughput, automated equipment. Given the current situation of the epidemic, point-of-care testing (POCT) is advantageous in terms of its ease of use, greater approachability on the user's end, more timely detection, and comparable accuracy and sensitivity, which could reduce the testing load on central hospitals. POCT is thus conducive to daily epidemic control and achieving early detection and treatment. This paper summarises the latest research advances in POCT-based SARS-CoV-2 detection methods, compares three categories of commercially available products, i.e., nucleic acid tests, immunoassays and novel sensors, and proposes the expectations for the development of POCT-based SARS-CoV-2 detection including greater accessibility, higher sensitivity and lower costs.

Logic control of microfluidics with smart colloid.

We report the successful realization of a microfluidic chip with switching and corresponding inverting functionalities. The chips are identical logic control components incorporating a type of smart colloid, giant electrorheological fluid (GERF), which possesses reversible characteristics via a liquid-solid phase transition under external electric field. Two pairs of electrodes embedded on the sides of two microfluidic channels serve as signal input and output, respectively. One, located in the GERF micro-channel is used to control the flow status of GERF, while another one in the ither micro-fluidic channel is used to detect the signal generated with a passing-by droplet (defined as a signal droplet). Switching of the GERF from the suspended state (off-state) to the flowing state (on-state) or vice versa in the micro-channel is controlled by the appearance of signal droplets whenever they pass through the detection electrode. The output on-off signals can be easily demonstrated, clearly matching with GERF flow status. Our results show that such a logic switch is also a logic IF gate, while its inverter functions as a NOT gate.

Wax-bonding 3D microfluidic chips.

We report a simple, low-cost and detachable microfluidic chip incorporating easily accessible paper, glass slides or other polymer films as the chip materials along with adhesive wax as the recycling bonding material. We use a laser to cut through the paper or film to form patterns and then sandwich the paper and film between glass sheets or polymer membranes. The hot-melt adhesive wax can realize bridge bonding between various materials, for example, paper, polymethylmethacrylate (PMMA) film, glass sheets, or metal plate. The bonding process is reversible and the wax is reusable through a melting and cooling process. With this process, a three-dimensional (3D) microfluidic chip is achievable by vacuating and venting the chip in a hot-water bath. To study the biocompatibility and applicability of the wax-based microfluidic chip, we tested the PCR compatibility with the chip materials first. Then we applied the wax-paper based microfluidic chip to HeLa cell electroporation (EP). Subsequently, a prototype of a 5-layer 3D chip was fabricated by multilayer wax bonding. To check the sealing ability and the durability of the chip, green fluorescence protein (GFP) recombinant Escherichia coli (E. coli) bacteria were cultured, with which the chemotaxis of E. coli was studied in order to determine the influence of antibiotic ciprofloxacin concentration on the E. coli migration.

Effect of additives on the growth of HKUST-1 crystals synthesized by microfluidic chips with concentration gradient.

Metal-organic frameworks (MOFs) have attracted considerable attention as novel nanoporous materials that combine the properties of organic and inorganic porous materials. HKUST-1 is one of the most well-developed and representative MOFs with wide applications in gas storage and separation, adsorption, and capture. In this study, we used microfluidics, an advanced technique of manipulation of small fluid volumes in microscale or even nanoscale channels, to investigate the effect of sodium dodecyl sulfate (SDS) on the growth of HKUST-1 crystals. We directly observed the morphological evolution of HKUST-1 crystals through droplet arrays with the SDS concentration gradient. The morphology of HKUST-1 evolved from cubic to cuboctahedron and finally to octahedron with increasing SDS concentration. The study results demonstrated the important role played by anions in solution in the structural regulation of HKUST-1.

Manipulation of microfluidic droplets by electrorheological fluid.

Microfluidics, especially droplet microfluidics, attracts more and more researchers from diverse fields, because it requires fewer materials and less time, produces less waste and has the potential of highly integrated and computer-controlled reaction processes for chemistry and biology. Electrorheological fluid, especially giant electrorheological fluid (GERF), which is considered as a kind of smart material, has been applied to the microfluidic systems to achieve active and precise control of fluid by electrical signal. In this review article, we will introduce recent results of microfluidic droplet manipulation, GERF and some pertinent achievements by introducing GERF into microfluidic system: digital generation, manipulation of "smart droplets" and droplet manipulation by GERF. Once it is combined with real-time detection, integrated chip with multiple functions can be realized.

An Automated and Miniaturized Rotating-Disk Device for Rapid Nucleic Acid Extraction.

The result of molecular diagnostic and detection greatly dependent on the quality and integrity of the isolated nucleic acid. In this work, we developed an automated miniaturized nucleic acid extraction device based on magnetic beads method, consisting of four components including a sample processing disc and its associated rotary power output mechanism, a pipetting module, a magnet module and an external central controller to enable a customizable and automated robust nucleic acid sample preparation. The extracted nucleic acid using 293T cells were verified using real-time polymerase chain reaction (PCR) and the data implies a comparable efficiency to a manual process, with the advantages of performing a flexible, time-saving (~10 min), and simple nucleic acid sample preparation.

Peptide-templated multifunctional nanoprobe for feasible electrochemical assay of intracellular kinase.

Protein kinases play a critical role in regulation of intracellular signal transduction, whose aberrant expression is closely associated with various dangerous human diseases. In this paper, we propose a feasible electrochemical assay of intracellular kinase by incorporating peptide nanoprobe-assisted signal labeling and signal amplification. Protein kinase A (PKA)-specific peptide P1 is self-assembled on the surface of a gold electrode, serine of which could be phosphorylated with catalysis of PKA in the presence of adenosine-5'-triphosphate (ATP). Another artificial peptide P2 contains a short template for preparation of copper nanoparticles-based nanoprobe (P2-CuNPs) and provides arginine residues for specific recognition of phosphorylation site. After PKA-catalyzed phosphorylation, phosphorylated P1 specially binds with P2-CuNPs through ultra-stable phosphate-guanidine interaction, and thus results in amplified electrochemical response from surface-attached CuNPs. Our method demonstrates a satisfactory sensitivity toward PKA detection with a detection limit of 0.0019 U/mL, which is also successfully applied in intracellular PKA assay and inhibitory study with high specificity comparable to ELISA. Therefore, the facile method suggests a promising potential use in kinase-related biochemical fundamental research, disease diagnosis and drug discovery in the future.

Synthesis of Biomaterials Utilizing Microfluidic Technology.

Recently, microfluidic technologies have attracted an enormous amount of interest as potential new tools for a large range of applications including materials synthesis, chemical and biological detection, drug delivery and screening, point-of-care diagnostics, and in-the-field analysis. Their ability to handle extremely small volumes of fluids is accompanied by additional benefits, most notably, rapid and efficient mass and heat transfer. In addition, reactions performed within microfluidic systems are highly controlled, meaning that many advanced materials, with uniform and bespoke properties, can be synthesized in a direct and rapid manner. In this review, we discuss the utility of microfluidic systems in the synthesis of materials for a variety of biological applications. Such materials include microparticles or microcapsules for drug delivery, nanoscale materials for medicine or cellular assays, and micro- or nanofibers for tissue engineering.

Preparation of barium strontium titanate Ba(1-x)Sr(x)TiO(3) (0

An ultrasonic irradiation method was applied to the sol/gel synthesis of the single-crystal cubic barium strontium titanate Ba(1-x)Sr(x)TiO(3) (0