Investigation of tumour environments through advancements in microtechnology and nanotechnology.

Cancer has a significant negative social and economic impact on both developed and developing countries. As a result, understanding the onset and progression of cancer is critical for developing therapies that can improve the well-being and health of individuals with cancer. With time, study has revealed, the tumor microenvironment has great influence on this process. Micro and nanoscale engineering techniques can be used to study the tumor microenvironment. Nanoscale and Microscale engineering use Novel technologies and designs with small dimensions to recreate the TME. Knowing how cancer cells interact with one another can help researchers develop therapeutic approaches that anticipate and counteract cancer cells' techniques for evading detection and fighting anti-cancer treatments, such as microfabrication techniques, microfluidic devices, nanosensors, and nanodevices used to study or recreate the tumor microenvironment. Nevertheless, a complicated action just like the growth and in cancer advancement, and their intensive association along the environment around it that has to be studied in more detail.

A high-throughput microfabricated platform for rapid quantification of metastatic potential.

Assays that measure morphology, proliferation, motility, deformability, and migration are used to study the invasiveness of cancer cells. However, native invasive potential of cells may be hidden from these contextual metrics because they depend on culture conditions. We created a micropatterned chip that mimics the native environmental conditions, quantifies the invasive potential of tumor cells, and improves our understanding of the malignancy signatures. Unlike conventional assays, which rely on indirect measurements of metastatic potential, our method uses three-dimensional microchannels to measure the basal native invasiveness without chemoattractants or microfluidics. No change in cell death or proliferation is observed on our chips. Using six cancer cell lines, we show that our system is more sensitive than other motility-based assays, measures of nuclear deformability, or cell morphometrics. In addition to quantifying metastatic potential, our platform can distinguish between motility and invasiveness, help study molecular mechanisms of invasion, and screen for targeted therapeutics.

Collection of microrobots for gentle cell manipulation.

Optically actuated soft microrobotic tools were designed for cell transportation, manipulation, and cell-to-cell interactions.

Microfabricated dynamic brain organoid cocultures to assess the effects of surface geometry on assembloid formation.

Organoids have emerged as valuable tools for the study of development and disease. Assembloids are formed by integrating multiple organoid types to create more complex models. However, the process by which organoids integrate to form assembloids remains unclear and may play an important role in the resulting organoid structure. Here, a microfluidic platform is developed that allows separate culture of distinct organoid types and provides the capacity to partially control the geometry of the resulting organoid surfaces. Removal of a microfabricated barrier then allows the shaped and positioned organoids to interact and form an assembloid. When midbrain and unguided brain organoids were allowed to assemble with a defined spacing between them, axonal projections from midbrain organoids and cell migration out of unguided organoids were observed and quantitatively measured as the two types of organoids fused together. Axonal projection directions were statistically biased toward other midbrain organoids, and unguided organoid surface geometry was found to affect cell invasion. This platform provides a tool to observe cellular interactions between organoid surfaces that are spaced apart in a controlled manner, and may ultimately have value in exploring neuronal migration, axon targeting, and assembloid formation mechanisms.

Analyzing the Micro-topography Guidance of Dictyostelium Cells Using Microfabricated Structures.

Over the last decade, the use of microfabricated substrates has proven pivotal for studying the effect of substrate topography on cell deformation and migration. Microfabrication techniques allow one to construct a transparent substrate with topographic features with high designability and reproducibility and thus well suited to experiments that microscopically address how spatial and directional bias are brought about in the cytoskeletal machineries and hence cell motility. While much of the progress in this avenue of study has so far been made in adhesive cells of epithelial and mesenchymal nature, whether related phenomena exist in less adhesive fast migrating cells is relatively unknown. In this chapter, we describe a method that makes use of micrometer-scale ridges to study fast-migrating Dictyostelium cells where it was recently shown that membrane evagination associated with macropinocytic cup formation plays a pivotal role in the topography sensing. The method requires only basic photolithography, and thus the step-by-step protocol should be a good entry point for cell biologists looking to incorporate similar microfabrication approaches.

Fly Me to the Micron: Microtechnologies for Drosophila Research.

Multicellular model organisms, such as Drosophila melanogaster (fruit fly), are frequently used in a myriad of biological research studies due to their biological significance and global standardization. However, traditional tools used in these studies generally require manual handling, subjective phenotyping, and bulk treatment of the organisms, resulting in laborious experimental protocols with limited accuracy. Advancements in microtechnology over the course of the last two decades have allowed researchers to develop automated, high-throughput, and multifunctional experimental tools that enable novel experimental paradigms that would not be possible otherwise. We discuss recent advances in microtechnological systems developed for small model organisms using D. melanogaster as an example. We critically analyze the state of the field by comparing the systems produced for different applications. Additionally, we suggest design guidelines, operational tips, and new research directions based on the technical and knowledge gaps in the literature. This review aims to foster interdisciplinary work by helping engineers to familiarize themselves with model organisms while presenting the most recent advances in microengineering strategies to biologists.

Mechanical Strain Induces and Increases Vesicular Release Monitored by Microfabricated Stretchable Electrodes.

Exocytosis involving the fusion of intracellular vesicles with cell membrane, is thought to be modulated by the mechanical cues in the microenvironment. Single-cell electrochemistry can offer unique information about the quantification and kinetics of exocytotic events; however, the effects of mechanical force on vesicular release have been poorly explored. Herein, we developed a stretchable microelectrode with excellent electrochemical stability under mechanical deformation by microfabrication of functionalized poly(3,4-ethylenedioxythiophene) conductive ink, which achieved real-time quantitation of strain-induced vesicular exocytosis from a single cell for the first time. We found that mechanical strain could cause calcium influx via the activation of Piezo1 channels in chromaffin cell, initiating the vesicular exocytosis process. Interestingly, mechanical strain increases the amount of catecholamines released by accelerating the opening and prolonging the closing of fusion pore during exocytosis. This work is expected to provide revealing insights into the regulatory effects of mechanical stimuli on vesicular exocytosis.

Cerenkov free micro-dosimetry in small-field radiation therapy technique.

Objective. Optical fiber-based scintillating dosimetry is a recent promising technique owing to the miniature size dosimeter and quality measurement in modern radiation therapy treatment. Despite several advantages, the major issue of using scintillating dosimeters is the Cerenkov effect and predominantly requires extra measurement corrections. Therefore, this work highlighted a novel micro-dosimetry technique to ensure Cerenkov-free measurement in radiation therapy treatment protocol by investigating several dosimetric characteristics.Approach.A micro-dosimetry technique was proposed with the performance evaluation of a novel infrared inorganic scintillator detector (IR-ISD). The detector essentially consists of a micro-scintillating head based on IR-emitting micro-clusters with a sensitive volume of 1.5 × 10-6mm3. The proposed system was evaluated under the 6 MV LINAC beam used in patient treatment. Overall measurements were performed using IBATMwater tank phantoms by following TRS-398 protocol for radiotherapy. Cerenkov measurements were performed for different small fields from 0.5 × 0.5 cm2to 10 × 10 cm2under LINAC. In addition, several dosimetric parameters such as percentage depth dose (PDD), high lateral resolution beam profiling, dose linearity, dose rate linearity, repeatability, reproducibility, and field output factor were investigated to realize the performance of the novel detector.Main results. This study highlighted a complete removal of the Cerenkov effect using a point-like miniature detector, especially for small field radiation therapy treatment. Measurements demonstrated that IR-ISD has acceptable behavior with dose rate variability (maximum standard deviation ∼0.18%) for the dose rate of 20-1000 cGy s-1. An entire linear response (R2= 1) was obtained for the dose delivered within the range of 4-1000 cGy, using a selected field size of 1 × 1 cm2. Perfect repeatability (max 0.06% variation from average) with day-to-day reproducibility (0.10% average variation) was observed. PDD profiles obtained in the water tank present almost identical behavior to the reference dosimeter with a build-up maximum depth dose at 1.5 cm. The small field of 0.5 × 0.5 cm2profiles have been characterized with a high lateral resolution of 100µm.Significance. Unlike recent plastic scintillation detector systems, the proposed micro-dosimetry system in this study requires no Cerenkov corrections and showed efficient performance for several dosimetric parameters. Therefore, it is expected that considering the detector correction factors, the IR-ISD system can be a suitable dose measurement tool, such as in small-field dose measurements, high and low gradient dose verification, and, by extension, in microbeam radiation and FLASH radiation therapy.

Thin-Film Piezoelectric Micromachined Ultrasound Transducers in Biomedical Applications: A Review.

Thin-film piezoelectric micromachined ultrasound transducers (PMUTs) are an increasingly relevant and well-researched field, and their biomedical importance has been growing as the technology continues to mature. This review article briefly discusses their history in biomedical use, provides a simple explanation of their principles for newer readers, and sheds light on the materials selection for these devices. Primarily, it discusses the significant applications of PMUTs in the biomedical industry and showcases recent progress that has been made in each application. The biomedical applications covered include common historical uses of ultrasound such as ultrasound imaging, ultrasound therapy, and fluid sensing, but additionally new and upcoming applications such as drug delivery, photoacoustic imaging, thermoacoustic imaging, biometrics, and intrabody communication. By including a device comparison chart for different applications, this review aims to assist microelectromechanical systems (MEMS) designers that work with PMUTs by providing a benchmark for recent research works. Furthermore, it puts forth a discussion on the current challenges being faced by PMUTs in the biomedical field, current and likely future research trends, and opportunities for PMUT development areas, as well as sharing the opinions and predictions of the authors on the state of this technology as a whole. The review aims to be a comprehensive introduction to these topics without diving excessively deep into existing literature.

A review of the recent achievements and future trends on 3D printed microfluidic devices for bioanalytical applications.

3D printing has revolutionized the manufacturing process of microanalytical devices by enabling the automated production of customized objects. This technology promises to become a fundamental tool, accelerating investigations in critical areas of health, food, and environmental sciences. This microfabrication technology can be easily disseminated among users to produce further and provide analytical data to an interconnected network towards the Internet of Things, as 3D printers enable automated, reproducible, low-cost, and easy fabrication of microanalytical devices in a single step. New functional materials are being investigated for one-step fabrication of highly complex 3D printed parts using photocurable resins. However, they are not yet widely used to fabricate microfluidic devices. This is likely the critical step towards easy and automated fabrication of sophisticated, complex, and functional 3D-printed microchips. Accordingly, this review covers recent advances in the development of 3D-printed microfluidic devices for point-of-care (POC) or bioanalytical applications such as nucleic acid amplification assays, immunoassays, cell and biomarker analysis and organs-on-a-chip. Finally, we discuss the future implications of this technology and highlight the challenges in researching and developing appropriate materials and manufacturing techniques to enable the production of 3D-printed microfluidic analytical devices in a single step.

Low-cost, versatile, and highly reproducible microfabrication pipeline to generate 3D-printed customised cell culture devices with complex designs.

Cell culture devices, such as microwells and microfluidic chips, are designed to increase the complexity of cell-based models while retaining control over culture conditions and have become indispensable platforms for biological systems modelling. From microtopography, microwells, plating devices, and microfluidic systems to larger constructs such as live imaging chamber slides, a wide variety of culture devices with different geometries have become indispensable in biology laboratories. However, while their application in biological projects is increasing exponentially, due to a combination of the techniques, equipment and tools required for their manufacture, and the expertise necessary, biological and biomedical labs tend more often to rely on already made devices. Indeed, commercially developed devices are available for a variety of applications but are often costly and, importantly, lack the potential for customisation by each individual lab. The last point is quite crucial, as often experiments in wet labs are adapted to whichever design is already available rather than designing and fabricating custom systems that perfectly fit the biological question. This combination of factors still restricts widespread application of microfabricated custom devices in most biological wet labs. Capitalising on recent advances in bioengineering and microfabrication aimed at solving these issues, and taking advantage of low-cost, high-resolution desktop resin 3D printers combined with PDMS soft lithography, we have developed an optimised a low-cost and highly reproducible microfabrication pipeline. This is thought specifically for biomedical and biological wet labs with not prior experience in the field, which will enable them to generate a wide variety of customisable devices for cell culture and tissue engineering in an easy, fast reproducible way for a fraction of the cost of conventional microfabrication or commercial alternatives. This protocol is designed specifically to be a resource for biological labs with limited expertise in those techniques and enables the manufacture of complex devices across the µm to cm scale. We provide a ready-to-go pipeline for the efficient treatment of resin-based 3D-printed constructs for PDMS curing, using a combination of polymerisation steps, washes, and surface treatments. Together with the extensive characterisation of the fabrication pipeline, we show the utilisation of this system to a variety of applications and use cases relevant to biological experiments, ranging from micro topographies for cell alignments to complex multipart hydrogel culturing systems. This methodology can be easily adopted by any wet lab, irrespective of prior expertise or resource availability and will enable the wide adoption of tailored microfabricated devices across many fields of biology.

Microfabrication of engineered Lactococcus lactis biocarriers with genetically programmed immunorecognition probes for sensitive lateral flow immunoassay of antibiotic in milk and lake water.

Micro/nanomaterials display considerable potential for increasing the sensitivity of lateral flow immunoassay (LFIA) by acting as 3D carriers for both antibodies and signals. The key to achieving high detection sensitivity depends on the probe's orientation on the material surface and its multivalent biomolecular interactions with targets. Here, we engineer Lactococcus lactis as the bacterial microcarrier (BMC) for a multivalent immunorecognition probe that was genetically programmed to display multifunctional components including a phage-screened single-chain variable fragment (scFv), an enhanced green fluorescent protein (eGFP), and a C-terminal peptidoglycan-binding domain (AcmA) anchored on BMC through the cell wall peptidoglycan. The innovative design of this biocarrier system, which incorporates a lab-on-a-chip microfluidic device, allows for the rapid and non-destructive self-assembly of the multivalent scFv-eGFP-AcmA@BMC probe, in which the 3D structure of BMC with a large peptidoglycan surface area facilitates the precisely orientated attachment and immobilization of scFv-eGFP-AcmA. This leads to a remarkable fluorescence aggregation amplification effect in LFIA, outperforming a monovalent 2D scFv-eGFP-AcmA probe for florfenicol detection. By designing a portable sensing device, we achieved an exceptionally low detection limit of 0.28 pg/mL and 0.21 pg/mL for florfenicol in lake water and milk sample, respectively. The successful microfabrication of this biocarrier holds potential to inspire innovative biohybrid designs for environment and food safety biosensing applications.

Highly Localized Chemical Sampling at Subsecond Temporal Resolution Enabled with a Silicon Nanodialysis Platform at Nanoliter per Minute Flows.

Microdialysis (MD) is a versatile and powerful technique for chemical profiling of biological tissues and is widely used for quantification of neurotransmitters, neuropeptides, metabolites, biomarkers, and drugs in the central nervous system as well as in dermatology, ophthalmology, and pain research. However, MD performance is severely limited by fundamental tradeoffs between chemical sensitivity, spatial resolution, and temporal response. Here, by using wafer-scale silicon microfabrication, we develop and demonstrate a nanodialysis (ND) sampling probe that enables highly localized chemical sampling with 100 µm spatial resolution and subsecond temporal resolution at high recovery rates. These performance metrics, which are 100-1000× superior to existing MD approaches, are enabled by a 100× reduction of the microfluidic channel cross-section, a corresponding drastic 100× reduction of flow rates to exceedingly slow few nL/min flows, and integration of a nanometer-thin nanoporous membrane with high transport flux into the probe sampling area. Miniaturized ND probes may allow for the minimally invasive and highly localized sampling and chemical profiling in live biological tissues with high spatiotemporal resolution for clinical, biomedical, and pharmaceutical applications.

Roadmap for Clinical Translation of Mobile Microrobotics.

Medical microrobotics is an emerging field to revolutionize clinical applications in diagnostics and therapeutics of various diseases. On the other hand, the mobile microrobotics field has important obstacles to pass before clinical translation. This article focuses on these challenges and provides a roadmap of medical microrobots to enable their clinical use. From the concept of a "magic bullet" to the physicochemical interactions of microrobots in complex biological environments in medical applications, there are several translational steps to consider. Clinical translation of mobile microrobots is only possible with a close collaboration between clinical experts and microrobotics researchers to address the technical challenges in microfabrication, safety, and imaging. The clinical application potential can be materialized by designing microrobots that can solve the current main challenges, such as actuation limitations, material stability, and imaging constraints. The strengths and weaknesses of the current progress in the microrobotics field are discussed and a roadmap for their clinical applications in the near future is outlined.

Microfabrication-based engineering of biomimetic dentin-like constructs to simulate dental aging.

Human dentin is a highly organized dental tissue displaying a complex microarchitecture consisting of micrometer-sized tubules encased in a mineralized type-I collagen matrix. As such, it serves as an important substrate for the adhesion of microbial colonizers and oral biofilm formation in the context of dental caries disease, including root caries in the elderly. Despite this issue, there remains a current lack of effective biomimetic in vitro dentin models that facilitate the study of oral microbial adhesion by considering the surface architecture at the micro- and nanoscales. Therefore, the aim of this study was to develop a novel in vitro microfabricated biomimetic dentin surface that simulates the complex surface microarchitecture of exposed dentin. For this, a combination of soft lithography microfabrication and biomaterial science approaches were employed to construct a micropitted PDMS substrate functionalized with mineralized type-I collagen. These dentin analogs were subsequently glycated with methylglyoxal (MGO) to simulate dentin matrix aging in vitro and analyzed utilizing an interdisciplinary array of techniques including atomic force microscopy (AFM), elemental analysis, and electron microscopy. AFM force-mapping demonstrated that the nanomechanical properties of the biomimetic constructs were within the expected biological parameters, and that mineralization was mostly predominated by hydroxyapatite deposition. Finally, dual-species biofilms of Streptococcus mutans and Candida albicans were grown and characterized on the biofunctionalized PDMS microchips, demonstrating biofilm-specific morphologic characteristics and confirming the suitability of this model for the study of early biofilm formation under controlled conditions. Overall, we expect that this novel biomimetic dentin model could serve as an in vitro platform to study oral biofilm formation or dentin-biomaterial bonding in the laboratory without the need for animal or human tooth samples in the future.

Breaking Barriers: Exploring Neurotransmitters through In Vivo vs. In Vitro Rivalry.

Neurotransmitter analysis plays a pivotal role in diagnosing and managing neurodegenerative diseases, often characterized by disturbances in neurotransmitter systems. However, prevailing methods for quantifying neurotransmitters involve invasive procedures or require bulky imaging equipment, therefore restricting accessibility and posing potential risks to patients. The innovation of compact, in vivo instruments for neurotransmission analysis holds the potential to reshape disease management. This innovation can facilitate non-invasive and uninterrupted monitoring of neurotransmitter levels and their activity. Recent strides in microfabrication have led to the emergence of diminutive instruments that also find applicability in in vitro investigations. By harnessing the synergistic potential of microfluidics, micro-optics, and microelectronics, this nascent realm of research holds substantial promise. This review offers an overarching view of the current neurotransmitter sensing techniques, the advances towards in vitro microsensors tailored for monitoring neurotransmission, and the state-of-the-art fabrication techniques that can be used to fabricate those microsensors.

Microfabrication of Gelatin Methacrylate/Hydroxyapatite Composites by Utilizing Alternate Soaking Process.

To construct a complex three-dimensional (3D) structure mimicking bone microstructure, hydrogel models of polymerized gelatin methacrylate (pGelMA) were fabricated by using stereolithography and modified with hydroxyapatite (HAp) via an alternate soaking process (ASP) using a solution of calcium and phosphate ions. Fabricated pGelMA line models whose widths were designed as 100, 300, and 600 µm were modified with HAp by ASP by changing the immersion time and number of cycles. After ASP, all of the line models with widths of 100, 300, and 600 µm were successfully modified with HAp, and large amounts of HAp were covered with the fabricated models by increasing both the immersion time and the number of cycles in ASP. HAp was observed near the surface of the line model with a width of 600 µm after ASP at an immersion time of 10 s, while the entire model was modified with HAp using ASPs for longer immersion times. The adhesion and spread of mesenchymal stem cells (MSCs) on the pGelMA-HAp discs depended on the ASP conditions. Moreover, the HAp modification of 3D pyramid models without alteration of the microstructure was also conducted. This two-step fabrication method of first fabricating frameworks of hydrogel models by stereolithography and subsequently modifying the fabricated models with HAp will lead to the development of 3D cell culture systems to support bone grafts or to create biological niches, such as artificial bone marrow.

Recent advancements in Mg-based micromotors for biomedical and environmental applications.

Synthetic micro/nanomotors have attracted considerable attention due to their promising potential in the field of biomedicine. Despite their great potential, major micromotors require chemical fuels or complex devices to generate external physical fields for propulsion. Therefore, for future practical medical and environmental applications, Mg-based micromotors that exhibit water-powered movement and thus eliminate the need for toxic fuels, and that display optimal biocompatibility and biodegradability, are attracting attention. In this review, we summarized the recent microarchitectural design of Mg-based micromotors for biomedical applications. We also highlight the mechanism for realizing their water-powered motility. Furthermore, recent biomedical and environmental applications of Mg-based micromotors are introduced. We envision that advanced Mg-based micromotors will have a profound impact in biomedicine.

Transforming medical device biofilm control with surface treatment using microfabrication techniques.

Biofilm deposition on indwelling medical devices and implanted biomaterials is frequently attributed to the prevalence of resistant infections in humans. Further, the nature of persistent infections is widely believed to have a biofilm etiology. In this study, the wettability of commercially available indwelling medical devices was explored for the first time, and its effect on the formation of biofilm was determined in vitro. Surprisingly, all tested indwelling devices were found to be hydrophilic, with surface water contact angles ranging from 60° to 75°. First, we established a thriving Candida albicans biofilm growth at 24 hours. in YEPD at 30°C and 37°C plus serum in vitro at Cyclic olefin copolymer (COC) modified surface, which was subsequently confirmed via scanning electron microscopy, while their cellular metabolic function was assessed using the XTT cell viability assay. Surfaces with patterned wettability show that a contact angle of 110° (hydrophobic) inhibits C. albicans planktonic and biofilm formation completely compared to robust growth at a contact angle of 40° (hydrophilic). This finding may provide a novel antimicrobial strategy to prevent biofilm growth and antimicrobial resistance on indwelling devices and prosthetic implants. Overall, this study provides valuable insights into the surface characteristics of medical devices and their potential impact on biofilm formation, leading to the development of improved approaches to control and prevent microbial biofilms and re-infections.

The speed and acceleration of the ball carrier and tackler into contact during front-on tackles in rugby league.

The aim was to use a combination of video analysis and microtechnology (10 Hz global positioning system [GPS]) to quantify and compare the speed and acceleration of ball-carriers and tacklers during the pre-contact phase (contact - 0.5s) of the tackle event during rugby league match-play. Data were collected from 44 professional male rugby league players from two Super League clubs across two competitive matches. Tackle events were coded and subject to three stages of inclusion criteria to identify front-on tackles. 10 Hz GPS data was synchronised with video to extract the speed and acceleration of the ball-carrier and tackler into each front-on tackle (n = 214). Linear mixed effects models (effect size [ES], confidence intervals, p-values) compared differences. Overall, ball-carriers (4.73 ± 1.12 m∙s-1) had greater speed into front-on tackles than tacklers (2.82 ± 1.07 m∙s-1; ES = 1.69). Ball-carriers accelerated (0.67 ± 1.01 m∙s-2) into contact whilst tacklers decelerated (-1.26 ± 1.36 m∙s-2; ES = 1.74). Positional comparisons showed speed was greater during back vs. back (ES = 0.66) and back vs. forward (ES = 0.40) than forward vs. forward tackle events. Findings can be used to inform strategies to improve performance and player welfare.