Harmonic acoustics for dynamic and selective particle manipulation.

Precise and selective manipulation of colloids and biological cells has long been motivated by applications in materials science, physics and the life sciences. Here we introduce our harmonic acoustics for a non-contact, dynamic, selective (HANDS) particle manipulation platform, which enables the reversible assembly of colloidal crystals or cells via the modulation of acoustic trapping positions with subwavelength resolution. We compose Fourier-synthesized harmonic waves to create soft acoustic lattices and colloidal crystals without using surface treatment or modifying their material properties. We have achieved active control of the lattice constant to dynamically modulate the interparticle distance in a high-throughput (>100 pairs), precise, selective and reversible manner. Furthermore, we apply this HANDS platform to quantify the intercellular adhesion forces among various cancer cell lines. Our biocompatible HANDS platform provides a highly versatile particle manipulation method that can handle soft matter and measure the interaction forces between living cells with high sensitivity.

Advances in Microsphere-based Super-resolution Imaging.

Techniques to resolve images beyond the diffraction limit of light with a large field of view (FOV) are necessary to foster progress in various fields such as cell and molecular biology, biophysics, and nanotechnology, where nanoscale resolution is crucial for understanding the intricate details of large-scale molecular interactions. Although several means of achieving super-resolutions exist, they are often hindered by factors such as high costs, significant complexity, lengthy processing times, and the classical tradeoff between image resolution and FOV. Microsphere-based super-resolution imaging has emerged as a promising approach to address these limitations. In this review, we delve into the theoretical underpinnings of microsphere-based imaging and the associated photonic nanojet. This is followed by a comprehensive exploration of various microsphere-based imaging techniques, encompassing static imaging, mechanical scanning, optical scanning, and acoustofluidic scanning methodologies. This review concludes with a forward-looking perspective on the potential applications and future scientific directions of this innovative technology.

A self-assembled graphene oxide adjuvant induces both enhanced humoral and cellular immune responses in influenza vaccine.

Antiviral vaccine is essential for preventing and controlling virus spreading, along with declining morbidity and mortality. A major challenge in effective vaccination lies in the ability to enhance both the humoral and cellular immune responses by adjuvants. Herein, self-assembled nanoparticles based on graphene oxide quantum dots with components of carnosine, resiquimod and Zn2+ ions, namely ZnGC-R, are designed as a new adjuvant for influenza vaccine. With its high capability for antigen-loading, ZnGC-R enhances antigen utilization, improves DC recruitment, and activates antigen-presenting cells. Single cell analysis of lymphocytes after intramuscular vaccination revealed that ZnGC-R generated multifaceted immune responses. ZnGC-R stimulated robust CD4+CCR7loPD-1hi Tfh and durable CD8+CD44hiCD62L- TEM immune responses, and simultaneously promoted the proliferation of CD26+ germinal center B cells. Besides, ZnGC-R elicited 2.53-fold higher hemagglutination-inhibiting antibody than commercial-licensed aluminum salt adjuvant. ZnGC-R based vaccine induced 342% stronger IgG antibody responses compared with vaccines with inactivated virus alone, leading to 100% in vivo protection efficacy against the H1N1 influenza virus challenge.

Fabricating Wood-Like Textures on Multicurved 3D Printed Architectural Elements.

Desktop three-dimensional printing (3DP) with the fused filament fabrication technique is widely employed for the manufacture of small-scale horizontally layered elements with a uniform striated appearance. What remains a challenge is developing printing processes that can automate the construction of more complex large-scale elements with a distinct fluid surface esthetic for architectural design applications. To address this challenge, this research explores the three-dimensional (3D) printing of multicurved wood-plastic composite panels that have the appeal of natural timber. It compares six-axis robotic technology and its ability to rotate the axes to print smooth curved layers in complex shapes with a large-scale, gantry-style 3D printer that is predominantly used for creating fast, horizontally aligned linear prints typical of 3D printing toolpathing. The prototype test results demonstrate that both technologies can produce multicurved elements with a timber-like esthetic.

Profiling extracellular vesicle surface proteins with 10 µL peripheral plasma within 4 h.

Extracellular vesicle (EV) surface proteins, expressed by primary tumours, are important biomarkers for early cancer diagnosis. However, the detection of these EV proteins is complicated by their low abundance and interference from non-EV components in clinical samples. Herein, we present a MEmbrane-Specific Separation and two-step Cascade AmpLificatioN (MESS2CAN) strategy for direct detection of EV surface proteins within 4 h. MESS2CAN utilises novel lipid probes (long chains linked by PEG2K with biotin at one end, and DSPE at the other end) and streptavidin-coated magnetic beads, permitting a 49.6% EV recovery rate within 1 h. A dual amplification strategy with a primer exchange reaction (PER) cascaded by the Cas12a system then allows sensitive detection of the target protein at 10 EV particles per microliter. Using 4 cell lines and 90 clinical test samples, we demonstrate MESS2CAN for analysing HER2, EpCAM and EGFR expression on EVs derived from cells and patient plasma. MESS2CAN reports the desired specificity and sensitivity of EGFR (AUC = 0.98) and of HER2 (AUC = 1) for discriminating between HER2-positive breast cancer, triple-negative breast cancer and healthy donors. MESS2CAN is a pioneering method for highly sensitive in vitro EV diagnostics, applicable to clinical samples with trace amounts of EVs.

A solution to the biophysical fractionation of extracellular vesicles: Acoustic Nanoscale Separation via Wave-pillar Excitation Resonance (ANSWER).

High-precision isolation of small extracellular vesicles (sEVs) from biofluids is essential toward developing next-generation liquid biopsies and regenerative therapies. However, current methods of sEV separation require specialized equipment and time-consuming protocols and have difficulties producing highly pure subpopulations of sEVs. Here, we present Acoustic Nanoscale Separation via Wave-pillar Excitation Resonance (ANSWER), which allows single-step, rapid (<10 min), high-purity (>96% small exosomes, >80% exomeres) fractionation of sEV subpopulations from biofluids without the need for any sample preprocessing. Particles are iteratively deflected in a size-selective manner via an excitation resonance. This previously unidentified phenomenon generates patterns of virtual, tunable, pillar-like acoustic field in a fluid using surface acoustic waves. Highly precise sEV fractionation without the need for sample preprocessing or complex nanofabrication methods has been demonstrated using ANSWER, showing potential as a powerful tool that will enable more in-depth studies into the complexity, heterogeneity, and functionality of sEV subpopulations.

Biomimetic apposition compound eye fabricated using microfluidic-assisted 3D printing.

After half a billion years of evolution, arthropods have developed sophisticated compound eyes with extraordinary visual capabilities that have inspired the development of artificial compound eyes. However, the limited 2D nature of most traditional fabrication techniques makes it challenging to directly replicate these natural systems. Here, we present a biomimetic apposition compound eye fabricated using a microfluidic-assisted 3D-printing technique. Each microlens is connected to the bottom planar surface of the eye via intracorporal, zero-crosstalk refractive-index-matched waveguides to mimic the rhabdoms of a natural eye. Full-colour wide-angle panoramic views and position tracking of a point source are realized by placing the fabricated eye directly on top of a commercial imaging sensor. As a biomimetic analogue to naturally occurring compound eyes, the eye's full-colour 3D to 2D mapping capability has the potential to enable a wide variety of applications from improving endoscopic imaging to enhancing machine vision for facilitating human-robot interactions.

Acoustohydrodynamic tweezers via spatial arrangement of streaming vortices.

Acoustics-based tweezers provide a unique toolset for contactless, label-free, and precise manipulation of bioparticles and bioanalytes. Most acoustic tweezers rely on acoustic radiation forces; however, the accompanying acoustic streaming often generates unpredictable effects due to its nonlinear nature and high sensitivity to the three-dimensional boundary conditions. Here, we demonstrate acoustohydrodynamic tweezers, which generate stable, symmetric pairs of vortices to create hydrodynamic traps for object manipulation. These stable vortices enable predictable control of a flow field, which translates into controlled motion of droplets or particles on the operating surface. We built a programmable droplet-handling platform to demonstrate the basic functions of planar-omnidirectional droplet transport, merging droplets, and in situ mixing via a sequential cascade of biochemical reactions. Our acoustohydrodynamic tweezers enables improved control of acoustic streaming and demonstrates a previously unidentified method for contact-free manipulation of bioanalytes and digitalized liquid handling based on a compact and scalable functional unit.

Acoustofluidic rotational tweezing enables high-speed contactless morphological phenotyping of zebrafish larvae.

Modern biomedical research and preclinical pharmaceutical development rely heavily on the phenotyping of small vertebrate models for various diseases prior to human testing. In this article, we demonstrate an acoustofluidic rotational tweezing platform that enables contactless, high-speed, 3D multispectral imaging and digital reconstruction of zebrafish larvae for quantitative phenotypic analysis. The acoustic-induced polarized vortex streaming achieves contactless and rapid (~1 s/rotation) rotation of zebrafish larvae. This enables multispectral imaging of the zebrafish body and internal organs from different viewing perspectives. Moreover, we develop a 3D reconstruction pipeline that yields accurate 3D models based on the multi-view images for quantitative evaluation of basic morphological characteristics and advanced combinations of metrics. With its contactless nature and advantages in speed and automation, our acoustofluidic rotational tweezing system has the potential to be a valuable asset in numerous fields, especially for developmental biology, small molecule screening in biochemistry, and pre-clinical drug development in pharmacology.

Acoustofluidic centrifuge for nanoparticle enrichment and separation.

Liquid droplets have been studied for decades and have recently experienced renewed attention as a simplified model for numerous fascinating physical phenomena occurring on size scales from the cell nucleus to stellar black holes. Here, we present an acoustofluidic centrifugation technique that leverages an entanglement of acoustic wave actuation and the spin of a fluidic droplet to enable nanoparticle enrichment and separation. By combining acoustic streaming and droplet spinning, rapid (<1 min) nanoparticle concentration and size-based separation are achieved with a resolution sufficient to identify and isolate exosome subpopulations. The underlying physical mechanisms have been characterized both numerically and experimentally, and the ability to process biological samples (including DNA segments and exosome subpopulations) has been successfully demonstrated. Together, this acoustofluidic centrifuge overcomes existing limitations in the manipulation of nanoscale (<100 nm) bioparticles and can be valuable for various applications in the fields of biology, chemistry, engineering, material science, and medicine.

Erratum: "Microfluidic approaches for cell-based molecular diagnosis" [Biomicrofluidics, 12, 051501 (2018)].

[This corrects the article DOI: 10.1063/1.5030891.].

Correction: High-throughput cell focusing and separation via acoustofluidic tweezers.

Correction for 'High-throughput cell focusing and separation via acoustofluidic tweezers' by Mengxi Wu et al., Lab Chip, 2018, 18, 3003-3010, DOI: .

Generating multifunctional acoustic tweezers in Petri dishes for contactless, precise manipulation of bioparticles.

Acoustic tweezers are a promising technology for the biocompatible, precise manipulation of delicate bioparticles ranging from nanometer-sized exosomes to millimeter-sized zebrafish larva. However, their widespread usage is hindered by their low compatibility with the workflows in biological laboratories. Here, we present multifunctional acoustic tweezers that can manipulate bioparticles in a disposable Petri dish. Various functionalities including cell patterning, tissue engineering, concentrating particles, translating cells, stimulating cells, and cell lysis are demonstrated. Moreover, leaky surface acoustic wave-based holography is achieved by encoding required phases in electrode profiles of interdigitated transducers. This overcomes the frequency and resolution limits of previous holographic techniques to control three-dimensional acoustic beams in microscale. This study presents a favorable technique for noncontact and label-free manipulation of bioparticles in commonly used Petri dishes. It can be readily adopted by the biological and medical communities for cell studies, tissue generation, and regenerative medicine.

Acoustic streaming vortices enable contactless, digital control of droplets.

Advances in lab-on-a-chip technologies are driven by the pursuit of programmable microscale bioreactors or fluidic processors that mimic electronic functionality, scalability, and convenience. However, few fluidic mechanisms allow for basic logic operations on rewritable fluidic paths due to cross-contamination, which leads to random interference between "fluidic bits" or droplets. Here, we introduce a mechanism that allows for contact-free gating of individual droplets based on the scalable features of acoustic streaming vortices (ASVs). By shifting the hydrodynamic equilibrium positions inside interconnected ASVs with multitonal electrical signals, different functions such as controlling the routing and gating of droplets on rewritable fluidic paths are demonstrated with minimal biochemical cross-contamination. Electrical control of this ASV-based mechanism allows for unidirectional routing and active gating behaviors, which can potentially be scaled to functional fluidic processors that can regulate the flow of droplets in a manner similar to the current in transistor arrays.

Precision Printing of Customized Cylindrical Capsules with Multifunctional Layers for Oral Drug Delivery.

Advances in personalized medicine will require custom drug formulations and delivery mechanisms. Herein, we demonstrate a new type of personalized capsule comprising of printed concentric cylindrical layers with each layer having a distinctive functional drug component. Poly ε-caprolactone (PCL) with paracetamol (APAP) and chlorpheniramine maleate (CM), synergistic drugs commonly used to alleviate influenza symptoms, are printed as an inner layer and outer layer, respectively, via microscaled electrohydrodynamic (EHD) printing. Polyvinylpyrrolidone (PVP) nanofibers are embedded as interlayers between the two printed PCL-drug layers using electrospinning (ES) techniques. The complete concentric cylindrical capsule with a 6 mm inner diameter and 15 mm length can be swallowed for oral drug delivery. After dissolution of the PVP interlayer, the capsule separates in two, with inner and outer capsules for continuous drug dosing and targeting. Imaging was achieved using a 3T MRI system which allowed temporal observations of the targeted release through the incorporation of nanoparticles (Fe3O4). The morphology and structure, chemical composition, mechanical properties, and biocompatibility of the capsules were studied in vitro. In summary, this new type of custom printed and electrospun capsule that enabled component separation, targeted drug release may advance personalized medicine via multidrug oral delivery.

Microfluidic approaches for cell-based molecular diagnosis.

The search for next-generation biomarkers has enabled cell-based diagnostics in a number of disciplines ranging from oncology to pharmacogenetics. However, cell-based diagnostics are still far from clinical reality due to the complex assays and associated protocols which typically require cell isolation, lysis, DNA extraction, amplification, and detection steps. Leveraging recent advances in microfluidics, many biochemical assays have been translated onto microfluidic platforms. We have compared and summarized recent advances in modular approaches toward the realization of fully-integrated, cell-based molecular diagnostics for clinical and point-of-care applications.

Three-Dimensional Electrohydrodynamic Printing and Spinning of Flexible Composite Structures for Oral Multidrug Forms.

A simple method to rapidly customize and to also mass produce oral dosage forms is arguably a current bottleneck in the development of modern personalized medicine. Specifically, delayed-release mechanisms with well-controlled dosage profiles for combinations of traditional Chinese herbal extracts and Western medications are not well established. Herein, we demonstrate a novel multidrug-loaded membrane sandwich with structures infused with ibuprofen (IBU) and Ganoderma lucidum polysaccharide (GLP) using three-dimensional electrohydrodynamic printing and electrospinning techniques. The resulting flexible membrane consists of microscaled, multilayered cellulose acetate (CA) membranes loaded with IBU in the shape of either concentric squares or circles, as the top and bottom layers of a sandwich structure. In between the CA-IBU layers are randomly electrospun polyvinyl pyrrolidone (PVP) layers loaded with GLP. The complete fibrous membrane sandwich can be folded and embedded into a 0-size capsule to achieve oral compliance. Simulated in vitro testing of gastric and intestinal fluids demonstrated a triphasic release profile. There was an immediate release of GLP after gastric juices dissolved the capsule shell and the PVP, followed by the short-term release of 60% of the IBU within an hour afterward, and the remaining IBU was released in a sustained manner following a Fickian diffusion profile. In summary, this multidrug (both hydrophilic and/or hydrophobic) oral system with precision-designed structures should enable personalized therapeutic dosing.

High-throughput cell focusing and separation via acoustofluidic tweezers.

Separation of particles and cells is an important function in many biological and biomedical protocols. Although a variety of microfluidic-based techniques have been developed so far, there is clearly still a demand for a precise, fast, and biocompatible method for separation of microparticles and cells. By combining acoustics and hydrodynamics, we have developed a method which we integrated into three-dimensional acoustofluidic tweezers (3D-AFT) to rapidly and efficiently separate microparticles and cells into multiple high-purity fractions. Compared with other acoustophoresis methods, this 3D-AFT method significantly increases the throughput by an order of magnitude, is label-free and gently handles the sorted cells. We demonstrate not only the separation of 10, 12, and 15 micron particles at a throughput up to 500 µl min-1 using this 3D-AFT method, but also the separation of erythrocytes, leukocytes, and cancer cells. This 3D-AFT method is able to meet various separation demands thus offering a viable alternative with potential for clinical applications.

Digital acoustofluidics enables contactless and programmable liquid handling.

For decades, scientists have pursued the goal of performing automated reactions in a compact fluid processor with minimal human intervention. Most advanced fluidic handling technologies (e.g., microfluidic chips and micro-well plates) lack fluid rewritability, and the associated benefits of multi-path routing and re-programmability, due to surface-adsorption-induced contamination on contacting structures. This limits their processing speed and the complexity of reaction test matrices. We present a contactless droplet transport and processing technique called digital acoustofluidics which dynamically manipulates droplets with volumes from 1 nL to 100 µL along any planar axis via acoustic-streaming-induced hydrodynamic traps, all in a contamination-free (lower than 10-10% diffusion into the fluorinated carrier oil layer) and biocompatible (99.2% cell viability) manner. Hence, digital acoustofluidics can execute reactions on overlapping, non-contaminated, fluidic paths and can scale to perform massive interaction matrices within a single device.

Label-free measurements of reaction kinetics using a droplet-based optofluidic device.

Label-free measurements of the reaction kinetics of a small sample volume are essential for efficient drug discovery, requiring methods and systems that are rapid, accurate, and cost-effective. Herein, we present an integrated optofluidic system for label-free characterization of reactions in a nanoliter reagent volume. This system contains a droplet-based microfluidic sampling section and an optical fiber-based spectroscopy detection section. By manipulating droplets containing reagents at certain concentrations at different times, quantifiable measurements via absorption spectroscopy can be made in a simple, sensitive, and high-throughput manner. We have demonstrated our system's capability by performing potency (IC50) assays of an inhibitor in a TEM-1 ß-lactamase (enzyme) and nitrocefin (substrate) system. This integrated platform can potentially provide an automated, label-free, and low-cost method for many other assays of reaction kinetics.