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Optics Express May 2024In this paper, a highly integrated terahertz (THz) biosensor is proposed and implemented, which pioneered the preparation of low-temperature gallium arsenide (LT-GaAs)...
In this paper, a highly integrated terahertz (THz) biosensor is proposed and implemented, which pioneered the preparation of low-temperature gallium arsenide (LT-GaAs) thin film photoconductive antenna (PCA) on the sensor for direct generation and detection of THz waves, simplifying complex terahertz time-domain spectroscopy (THz-TDS) systems. A latch type metasurface is deposited in the detection region to produce a resonance absorption peak at 0.6 THz that is independent of polarisation. Microfluidics is utilised and automatic injection is incorporated to mitigate the experimental effects of hydrogen bond absorption of THz waves in aqueous-based environment. Additionally, cell damage is minimised by regulating the cell flow rate. The biosensor was utilised to detect the concentration of three distinct sizes of bacteria with successful results. The assay was executed as a proof of concept to detect two distinct types of breast cancer cells. Based on the experimental findings, it has been observed that the amplitude and blueshift of the resonance absorption peaks have the ability to identify and differentiate various cancer cell types. The findings of this study introduce a novel approach for developing microfluidic THz metasurface biosensors that possess exceptional levels of integration, sensitivity, and rapid label-free detection capabilities.
Topics: Gallium; Arsenicals; Biosensing Techniques; Terahertz Spectroscopy; Humans; Equipment Design; Microfluidics
PubMed: 38858883
DOI: 10.1364/OE.518638 -
Microbial Cell Factories Jun 2024In vitro expression involves the utilization of the cellular transcription and translation machinery in an acellular context to produce one or more proteins of interest...
BACKGROUND
In vitro expression involves the utilization of the cellular transcription and translation machinery in an acellular context to produce one or more proteins of interest and has found widespread application in synthetic biology and in pharmaceutical biomanufacturing. Most in vitro expression systems available are active at moderate temperatures, but to screen large libraries of natural or artificial genetic diversity for highly thermostable enzymes or enzyme variants, it is instrumental to enable protein synthesis at high temperatures.
OBJECTIVES
Develop an in vitro expression system operating at high temperatures compatible with enzymatic assays and with technologies that enable ultrahigh-throughput protein expression in reduced volumes, such as microfluidic water-in-oil (w/o) droplets.
RESULTS
We produced cell-free extracts from Thermus thermophilus for in vitro translation including thermostable enzymatic cascades for energy regeneration and a moderately thermostable RNA polymerase for transcription, which ultimately limited the temperature of protein synthesis. The yield was comparable or superior to other thermostable in vitro expression systems, while the preparation procedure is much simpler and can be suited to different Thermus thermophilus strains. Furthermore, these extracts have enabled in vitro expression in microfluidic droplets at high temperatures for the first time.
CONCLUSIONS
Cell-free extracts from Thermus thermophilus represent a simpler alternative to heavily optimized or pure component thermostable in vitro expression systems. Moreover, due to their compatibility with droplet microfluidics and enzyme assays at high temperatures, the reported system represents a convenient gateway for enzyme screening at higher temperatures with ultrahigh-throughput.
Topics: Thermus thermophilus; Transcription, Genetic; Protein Biosynthesis; Microfluidics; Cell-Free System; DNA-Directed RNA Polymerases; Temperature; Hot Temperature; Bacterial Proteins
PubMed: 38858677
DOI: 10.1186/s12934-024-02440-y -
Scientific Reports Jun 2024Microfluidic devices with complex geometries and obstacles have attracted considerable interest in biomedical engineering and chemical analysis. Understanding droplet...
Microfluidic devices with complex geometries and obstacles have attracted considerable interest in biomedical engineering and chemical analysis. Understanding droplet breakup behavior within these systems is crucial for optimizing their design and performance. This study investigates the influence of triangular obstacles on droplet breakup processes in microchannels. Two distinct types of triangular obstructions, positioned at the bifurcation (case I) and aligned with the flow (case II), are analyzed to evaluate their impact on droplet behavior. The investigation considers various parameters, including the Capillary number (Ca), non-dimensional droplet length (L*), non-dimensional height (A*), and non-dimensional base length (B*) of the triangle. Utilizing numerical simulations with COMSOL software, the study reveals that the presence of triangular obstacles significantly alters droplet breakup dynamics. Importantly, the shape and location of the obstacle emerge as key factors governing breakup characteristics. Results indicate faster breakup of the initial droplet when the obstacle is positioned in the center of the microchannel for case I. For case II, the study aims to identify conditions under which droplets either break up into unequal-sized entities or remain intact, depending on various flow conditions. The findings identify five distinct regimes: no breakup, breakup without a tunnel, breakup with a tunnel, droplet fragmentation into unequal-sized parts, and sorting. These regimes depend on the presence or absence of triangular obstacles and the specific flow conditions. This investigation enhances our understanding of droplet behavior within intricate microfluidic systems and provides valuable insights for optimizing the design and functionality of droplet manipulation and separation devices. Notably, the results emphasize the significant role played by triangular obstacles in droplet breakup dynamics, with the obstacle's shape and position being critical determinants of breakup characteristics.
PubMed: 38858444
DOI: 10.1038/s41598-024-63922-y -
Scientific Reports Jun 2024We introduce magnetophoresis-based microfluidics for sorting biological targets using positive Magnetophoresis (pM) for magnetically labeled particles and negative...
We introduce magnetophoresis-based microfluidics for sorting biological targets using positive Magnetophoresis (pM) for magnetically labeled particles and negative Magnetophoresis (nM) for label-free particles. A single, externally magnetized ferromagnetic wire induces repulsive forces and is positioned across the focused sample flow near the main channel's closed end. We analyze magnetic attributes and separation performance under two transverse dual-mode magnetic configurations, examining magnetic fields, hydrodynamics, and forces on microparticles of varying sizes and properties. In pM, the dual-magnet arrangement (DMA) for sorting three distinct particles shows higher magnetic gradient generation and throughput than the single-magnet arrangement (SMA). In nM, the numerical results for SMA sorting of red blood cells (RBCs), white blood cells (WBCs), and prostate cancer cells (PC3-9) demonstrate superior magnetic properties and throughput compared to DMA. Magnetized wire linear movement is a key design parameter, allowing device customization. An automated device for handling more targets can be created by manipulating magnetophoretic repulsion forces. The transverse wire and magnet arrangement accommodate increased channel depth without sacrificing efficiency, yielding higher throughput than other devices. Experimental validation using soft lithography and 3D printing confirms successful sorting and separation, aligning well with numerical results. This demonstrates the successful sorting and separating of injected particles within a hydrodynamically focused sample in all systems. Both numerical and experimental findings indicate a separation accuracy of 100% across various Reynolds numbers. The primary channel dimensions measure 100 µm in height and 200 µm in width. N52 permanent magnets were employed in both numerical simulations and experiments. For numerical simulations, a remanent flux density of 1.48 T was utilized. In the experimental setup, magnets measuring 0.5 × 0.5 × 0.125 inches and 0.5 × 0.5 × 1 inch were employed. The experimental data confirm the device's capability to achieve 100% separation accuracy at a Reynolds number of 3. However, this study did not explore the potential impact of increased flow rates on separation accuracy.
Topics: Humans; Microfluidic Analytical Techniques; Cell Separation; Erythrocytes; Microfluidics; Leukocytes; Hydrodynamics; Cell Line, Tumor
PubMed: 38858424
DOI: 10.1038/s41598-024-64330-y -
Biomedical Optics Express May 2024Carbohydrates are pivotal biomolecules in biochemistry; this study employs terahertz time-domain spectroscopy (THz-TDS) to investigate the spectral characteristics of...
Carbohydrates are pivotal biomolecules in biochemistry; this study employs terahertz time-domain spectroscopy (THz-TDS) to investigate the spectral characteristics of trehalose and its hydrate across the 0.1 to 2.2 THz frequency range. Notable differences in spectra between the two compounds were observed. Density Functional Theory (DFT) simulations of the crystal structure were conducted to elucidate this phenomenon. The consistency between experimental results and simulations substantiates the reliability of the experimental findings. Additionally, the spectral characteristics of these carbohydrates in solution were examined using microfluidic chip technology. This approach facilitates a comprehensive comparison of their behaviors in both solid and solution states.
PubMed: 38855704
DOI: 10.1364/BOE.519006 -
Biomedical Optics Express May 2024Optofluidic devices hold great promise in biomedical diagnostics and testing because of their advantages of miniaturization, high sensitivity, high throughput, and high...
Optofluidic devices hold great promise in biomedical diagnostics and testing because of their advantages of miniaturization, high sensitivity, high throughput, and high scalability. However, conventional silicon-based photonic chips suffer from complicated fabrication processes and less flexibility in functionalization, thus hindering their development of cost-effective biomedical diagnostic devices for daily tests and massive applications in responding to public health crises. In this paper, we present an optofluidic chip based on directly printed polymer optical waveguide Mach-Zehnder interferometer (MZI) sensors for label-free biomarker detection. With digital ultraviolet lithography technology, high-sensitivity asymmetric MZI microsensors based on a width-tailored optical waveguide are directly printed and vertically integrated with a microfluidic layer to make an optofluidic chip. Experimental results show that the sensitivity of the directly printed polymer optical waveguide MZI sensor is about 1695.95 nm/RIU. After being modified with capture molecules, i.e., goat anti-human immunoglobulin G (IgG), the polymer optical waveguide MZI sensors can on-chip detect human IgG at the concentration level of 1.78 pM. Such a polymer optical waveguide-based optofluidic chip has the advantages of miniaturization, cost-effectiveness, high sensitivity, and ease in functionalization and thus has great potential in the development of daily available point-of-care diagnostic and testing devices.
PubMed: 38855677
DOI: 10.1364/BOE.523055 -
Frontiers in Bioengineering and... 2024Lung cancer is a malignant tumour with the highest incidence and mortality worldwide. Clinically effective therapy strategies are underutilized owing to the lack of... (Review)
Review
Lung cancer is a malignant tumour with the highest incidence and mortality worldwide. Clinically effective therapy strategies are underutilized owing to the lack of efficient models for evaluating drug response. One of the main reasons for failure of anticancer drug therapy is development of drug resistance. Anticancer drugs face severe challenges such as poor biodistribution, restricted solubility, inadequate absorption, and drug accumulation. In recent years, "organ-on-a-chip" platforms, which can directly regulate the microenvironment of biomechanics, biochemistry and pathophysiology, have been developed rapidly and have shown great potential in clinical drug research. Lung-on-a-chip (LOC) is a new 3D model of bionic lungs with physiological functions created by micromachining technology on microfluidic chips. This approach may be able to partially replace animal and 2D cell culture models. To overcome drug resistance, LOC realizes personalized prediction of drug response by simulating the lung-related microenvironment , significantly enhancing therapeutic effectiveness, bioavailability, and pharmacokinetics while minimizing side effects. In this review, we present an overview of recent advances in the preparation of LOC and contrast it with earlier models. Finally, we describe recent advances in LOC. The combination of this technology with nanomedicine will provide an accurate and reliable treatment for preclinical evaluation.
PubMed: 38854856
DOI: 10.3389/fbioe.2024.1378299 -
BioRxiv : the Preprint Server For... May 2024In nature, organisms experience combinations of stressors. However, laboratory studies typically simplify reality and focus on the effects of an individual stressor....
In nature, organisms experience combinations of stressors. However, laboratory studies typically simplify reality and focus on the effects of an individual stressor. Here, we use a microfluidic approach to simultaneously provide a physical stressor (shear flow) and a chemical stressor (H O ) to the human pathogen . By treating cells with levels of flow and H O that commonly co-occur in nature, we discover that previous reports significantly overestimate the H O levels required to block bacterial growth. Specifically, we establish that flow increases H O effectiveness 50-fold, explaining why previous studies lacking flow required much higher concentrations. Using natural H O levels, we identify the core H O regulon, characterize OxyR-mediated dynamic regulation, and dissect the redundant roles of multiple H O scavenging systems. By examining single-cell behavior, we serendipitously discover that the combined effects of H O and flow block pilus-driven surface migration. Thus, our results counter previous studies and reveal that natural levels of H O and flow synergize to restrict bacterial colonization and survival. By studying two stressors at once, our research highlights the limitations of oversimplifying nature and demonstrates that physical and chemical stress can combine to yield unpredictable effects.
PubMed: 38853869
DOI: 10.1101/2024.05.27.595753 -
Chinese Medicine Jun 2024Benefiting from the complex system composed of various constituents, medicament portions, species, and places of origin, traditional Chinese medicine (TCM) possesses... (Review)
Review
Benefiting from the complex system composed of various constituents, medicament portions, species, and places of origin, traditional Chinese medicine (TCM) possesses numerous customizable and adaptable efficacies in clinical practice guided by its theories. However, these unique features are also present challenges in areas such as quality control, screening active ingredients, studying cell and organ pharmacology, and characterizing the compatibility between different Chinese medicines. Drawing inspiration from the holistic concept, an integrated strategy and pattern more aligned with TCM research emerges, necessitating the integration of novel technology into TCM modernization. The microfluidic chip serves as a powerful platform for integrating technologies in chemistry, biology, and biophysics. Microfluidics has given rise to innovative patterns like lab-on-a-chip and organoids-on-a-chip, effectively challenging the conventional research paradigms of TCM. This review provides a systematic summary of the nature and advanced utilization of microfluidic chips in TCM, focusing on quality control, active ingredient screening/separation, pharmaceutical analysis, and pharmacological/toxicological assays. Drawing on these remarkable references, the challenges, opportunities, and future trends of microfluidic chips in TCM are also comprehensively discussed, providing valuable insights into the development of TCM.
PubMed: 38853247
DOI: 10.1186/s13020-024-00956-4 -
Current Opinion in Structural Biology Jun 2024Proteins execute numerous cell functions in concert with one another in protein-protein interactions (PPI). While essential in each cell, such interactions are not... (Review)
Review
Proteins execute numerous cell functions in concert with one another in protein-protein interactions (PPI). While essential in each cell, such interactions are not identical from cell to cell. Instead, PPI heterogeneity contributes to cellular phenotypic heterogeneity in health and diseases such as cancer. Understanding cellular phenotypic heterogeneity thus requires measurements of properties of PPIs such as abundance, stoichiometry, and kinetics at the single-cell level. Here, we review recent, exciting progress in single-cell PPI measurements. Novel technology in this area is enabled by microscale and microfluidic approaches that control analyte concentration in timescales needed to outpace PPI disassembly kinetics. We describe microscale innovations, needed technical capabilities, and methods poised to be adapted for single-cell analysis in the near future.
PubMed: 38848654
DOI: 10.1016/j.sbi.2024.102860