Acoustic separation and concentration of exosomes for nucleotide detection: ASCENDx

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Figure: Operating mechanism of the ASCENDx platform.

Efficient isolation and analysis of exosomal biomarkers hold transformative potential in biomedical applications. However, current methods are prone to contamination and require costly consumables, expensive equipment, and skilled personnel. Here, we introduce an innovative spaceship-like disc that allows Acoustic Separation and Concentration of Exosomes and Nucleotide Detection: ASCENDx. We created ASCENDx to use acoustically driven disc rotation on a spinning droplet to generate swift separation and concentration of exosomes from patient plasma samples. Integrated plasmonic nanostars on the ASCENDx disc enable label-free detection of enriched exosomes via surface-enhanced Raman scattering. Direct detection of circulating exosomal microRNA biomarkers from patient plasma samples by the ASCENDx platform facilitated a diagnostic assay for colorectal cancer with 95.8% sensitivity and 100% specificity. ASCENDx overcomes existing limitations in exosome-based molecular diagnostics and holds a powerful position for future biomedical research, precision medicine, and point-of-care medical diagnostics.

Cellular Immunity Analysis by Modular Acoustofluidic Platform: CIAMAP

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Figure: Schematic of CIAMAP-facilitated cell pairing and immunity mechanism exploration.

The study of molecular mechanisms at the single-cell level holds immense potential for enhancing immunotherapy and understanding neuroinflammation and neurodegenerative diseases by identifying previously concealed pathways within a diverse range of paired cells. However, existing single-cell pairing platforms have limitations in low pairing efficiency, complex manual operation procedures, and single-use functionality. Here, we report a multiparametric cellular immunity analysis by a modular acoustofluidic platform: CIAMAP. This platform enables users to efficiently sort and collect effector-target (i.e., NK92-K562) cell pairs and monitor the real-time dynamics of immunological response formation. Furthermore, we conducted transcriptional and protein expression analyses to evaluate the pathways that mediate effector cytotoxicity toward target cells, as well as the synergistic effect of doxorubicin on the cellular immune response. Our CIAMAP can provide promising building blocks for high-throughput quantitative single-cell level coculture to understand intercellular communication while also empowering immunotherapy by precision analysis of immunological synapses.

Acoustofluidic multimodal diagnostic system for Alzheimer's disease

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Figure: Schematic illustration of the acoustofluidics-based diagnostic system (ADx) for Alzheimer's disease. Liquid biopsy is employed to test AD patients' plasma samples for circulating protein biomarkers such as Aβ and Tau (A–B). An acoustofluidic separation chip is employed for removing contaminating bioparticles and isolating AD-specific biomarkers from patient plasma (C). An acoustofluidic microreactor (D) is employed for developing functional nanoarray-based SERS (E) and electrochemical immunosensor (F).

Alzheimer's disease (AD) is a progressive and irreversible neurodegenerative brain disorder that affects tens of millions of older adults worldwide and has significant economic and societal impacts. Despite its prevalence and severity, early diagnosis of AD remains a considerable challenge. Here we report an integrated acoustofluidics-based diagnostic system (ADx), which combines triple functions of acoustics, microfluidics, and orthogonal biosensors for clinically accurate, sensitive, and rapid detection of AD biomarkers from human plasma. We design and fabricate a surface acoustic wave-based acoustofluidic separation device to isolate and purify AD biomarkers to increase the signal-to-noise ratio. Multimodal biosensors within the integrated ADx are fabricated by in-situ patterning of the ZnO nanorod array and deposition of Ag nanoparticles onto the ZnO nanorods for surface-enhanced Raman scattering (SERS) and electrochemical immunosensors. We obtain the label-free detections of SERS and electrochemical immunoassay of clinical plasma samples from AD patients and healthy controls with high sensitivity and specificity. We believe that this efficient integration provides promising solutions for the early diagnosis of AD.

Acoustofluidic Interfaces for the Mechanobiological Secretome of MSCs

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Figure: a Schematic of the formation of MSC aggregates via acoustofluidic assembly and N-cadherin mediated cell–cell interactions to enhance the MSC secretome. b The simulation results showed that a three-dimensional circular vortex tube generated by a high-intensity focal point at the center of the substrate functioned as a virtual wall, trapping and aggregating cells at the center of the droplet. c Illustration of the comparison between 2D monolayer MSC culture and acoustofluidic 3D MSC aggregation. d Fluorescence images of MSCs before/after the acoustofluidic assembly, and the morphology of MSC aggregates after 3 days of incubation. The green color indicates alive cells. The blue color indicates cell nuclei, and the red color indicates the cytoskeleton. n = 4 tests with similar results. Scale bar: 100 μm. e CyQUANT™ Cell Proliferation Assay measuring the DNA contents of different groups. The statistical analysis was performed using one-way ANOVA with Tukey’s post-hoc test. Data are graphed as the mean ± SD (n = 4, biological repeats). Source data are provided as a Source Data file.

Acoustothermal transfection for cell therapy

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Figure: Acoustothermal transfection for enhancing cell membrane and nuclear envelope permeabilities.

Transfected stem cells and T cells are promising in personalized cell therapy and immunotherapy against various diseases. However, existing transfection techniques face a fundamental trade-off between transfection efficiency and cell viability; achieving both simultaneously remains a substantial challenge. This study presents an acoustothermal transfection method that leverages acoustic and thermal effects on cells to enhance the permeability of both the cell membrane and nuclear envelope to achieve safe, efficient, and high-throughput transfection of primary T cells and stem cells. With this method, two types of plasmids were simultaneously delivered into the nuclei of mesenchymal stem cells (MSCs) with efficiencies of 89.6 ± 1.2%. CXCR4-transfected MSCs could efficiently target cerebral ischemia sites in vivo and reduce the infarct volume in mice. Our acoustothermal transfection method addresses a key bottleneck in balancing the transfection efficiency and cell viability, which can become a powerful tool in the future for cellular and gene therapies.

Acoustoelectronic nanotweezers enable dynamic and large-scale control of nanomaterials

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Figure: a Acoustoelectronic fields are generated via dynamic acoustic wave interactions. These acoustic waves have minimal out-of-plane vibrations and associated acoustic attenuation losses in a fluid. F is the surface electric potential. b Schematic side-view of the electric field distribution and trapping positions for particles with different polarizabilities relative to the medium (red sphere: high polarizability; green sphere: low polarizability). c Schematic mechanism of AENT on manipulating nanoparticles with lower (p_np < p_m) or higher (p_np > p_m) polarizability than the medium in 3D space by tuning the phases and amplitudes of the acoustic waves. Δφ₁ indicates the phase variation of IDT₁. ΔA₁₂ indicates the amplitudes variation of IDT₁ and IDT₂. d Candidate excitation configurations based on nine potential single-crystal piezoelectric materials for AENT. κ_AET is the acoustoelectronic efficiency, which is defined as the ratio between the surface electric potential and the excitation voltage on the transducer in a standing wave mode. u_fluid is the acoustic streaming speed under consistent excitation amplitudes on different crystals. e Macroscopic materials with pre-designed nanotextures fabricated by AENT. Insets: microscopic images of PDMS films containing aligned carbon nanotubes and 100 nm PS beads, PEG hydrogels containing textured FITC-BSA proteins (66 kDa) and FITC-dextran (3 kDa). Scale bar: 60 μm.

The ability to precisely manipulate nano-objects on a large scale can enable the fabrication of materials and devices with tunable optical, electromagnetic, and mechanical properties. However, the dynamic, parallel manipulation of nanoscale colloids and materials remains a significant challenge. Here, we demonstrate acoustoelectronic nanotweezers, which combine the precision and robustness afforded by electronic tweezers with versatility and large-field dynamic control granted by acoustic tweezing techniques, to enable the massively parallel manipulation of sub-100 nm objects with excellent versatility and controllability. Using this approach, we demonstrated the complex patterning of various nanoparticles (e.g., DNAs, exosomes, ~3 nm graphene flakes, ~6 nm quantum dots, ~3.5 nm proteins, and ~1.4 nm dextran), fabricated macroscopic materials with nano-textures, and performed high-resolution, single nanoparticle manipulation. Various nanomanipulation functions, including transportation, concentration, orientation, pattern-overlaying, and sorting, have also been achieved using a simple device configuration. Altogether, acoustoelectronic nanotweezers overcome existing limitations in nano-manipulation and hold great potential for a variety of applications in the fields of electronics, optics, condensed matter physics, metamaterials, and biomedicine.

Digital acoustofluidics enables contactless and programmable liquid handling

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Figure: Digital acoustofluidics for contactless and programmable droplet manipulation. a Schematic showing one unit consisting of four IDTs (one pixel) in the digital acoustofluidic device. The four IDTs can be selectively excited to translate droplets along the ±x and ±y directions. The aqueous droplets are isolated from the piezoelectric substrate by an inert carrier fluid to prevent direct contact with surfaces. The IDT (bottom left) embedded beneath the carrier fluid generates SAWs that pumps out fluid in the ±y directions and pumps in fluid in the ±x directions. The red and blue droplets are separately trapped at the two symmetric hydrodynamic wells near the flanks of an IDT. The blue droplet is translated toward a well on the other side of the excited transducer. The reflux streamlines are shown in black. b A photo showing the digital acoustofluidic device with a drop of blood floating on the carrier layer of fluorinated oil

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.

Contactless, programmable acoustofluidic manipulation of objects on water  

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Figure: Contactless, programmable acoustofluidic manipulation of objects on water. (a) Schematic showing one unit of the acoustofluidic device. The oil droplets (red and blue spheres) are isolated in water to prevent direct contact with surfaces. This unit IDT has a hollow square structure and generates SAWs in ±x and ±y directions. The acoustic waves propagate, leak into the carrier liquid to form jets which propulse droplets along orthogonal axes on the surface of water. (b) Device image. The acoustofluidic chip consists of 36 pixels, each individually excitable.

Contact-free manipulation of small objects (e.g., cells, tissues, and droplets) using acoustic waves eliminates physical contact with structures and undesired surface adsorption. Pioneering acoustic-based, contact-free manipulation techniques (e.g., acoustic levitation) enable programmable manipulation but are limited by evaporation, bulky transducers, and inefficient acoustic coupling in air. Herein, we report an acoustofluidic mechanism for the contactless manipulation of small objects on water. A hollow-square-shaped interdigital transducer (IDT) is fabricated on lithium niobate (LiNbO₃), immersed in water and used as a sound source to generate acoustic waves and as a micropump to pump fluid in the ±x and ±y orthogonal directions. As a result, objects which float adjacent to the excited IDT can be pushed unidirectionally (horizontally) in ±x and ±y following the directed acoustic wave propagation. A fluidic processor was developed by patterning IDT units in a 6-by-6 array. We demonstrate contactless, programmable manipulation on water of oil droplets and zebrafish larvae. This acoustofluidic-based manipulation opens avenues for the contactless, programmable processing of materials and small biosamples.

Acoustic streaming vortices enable contactless, digital control of droplets

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Figure: ASV-based droplet manipulation. (A) Schematic showing a typical droplet processing unit. The droplets (i.e., red and blue spheres) over the transducers are guided into the center between the barrel-like ASVs (labeled as Vortices) following the recirculating inflow. These droplets are unidirectionally routed along the linear array of IDTs by shifting the sequence of working frequencies. The portions of the IDTs shaded in purple (e.g., IDT_i) indicate that they are being excited by a high-amplitude signal with frequency f_i. (B) The photo shows a bifurcated device with a particle floating above the transparent carrier oil layer. (C) The general control schematic for the droplet processing unit. The unit is composed of K + M + N interconnected IDTs [denoted as IDTkI, IDTmL, and IDTnR (k = 1,2, …, K; m = 1,2, …, M; n = 1,2, …, N)] with tuned working frequencies. Multitonal signals (i.e., S_IL or S_IR) encoded with a series of different frequencies, amplitudes, durations, and initiation times (i.e., [fmL,AmL,TmL,tmL]K+M or [fnR,AnR,TnR,tnR]K+N) are the excitation signals into the droplet processing unit and can direct the droplet from IDT1I to the left or right port, respectively. The blue shaded area indicates the resulting virtual bifurcated channel for droplet translation. Photo credits: Peiran Zhang, Duke University.

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.

 Acoustofluidic devices controlled by cell-phone

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Figure (a) Comparison between conventional acoustofluidic device operation and portable operation. The portable control platform can be used with either a traditional benchtop microscope or a portable microscope based on a cell phone camera. Not to scale. (b) Photo of the cell phone, modified speaker and acoustofluidic device. Signal measured from the speaker (c) before and (d) after passing through the RLC filter circuit to increase the voltage.

Here we report on the effort to provide an easy-to-use and portable system for controlling sharp-edge-based acoustofluidic devices. With the use of a cell phone and a modified Bluetooth® speaker, on-demand and hands-free pumping and mixing are achieved. Additionally, a novel design for a sharp-edge-based acoustofluidic device is proposed that combines both pumping and mixing functions into a single device, thus removing the need for external equipment typically needed to accomplish these two tasks. These applications serve to demonstrate the potential function that acoustofluidic devices can provide in point-of-care platforms. 

Reference:

1. Bachman, H., Huang, P.H., Zhao, S.G., Yang, S.Y., Zhang, P.R., Fu, H., and Huang, T.J., Acoustofluidic Devices Controlled by Cell Phones, Lab on a Chip, Vol. 18, pp. 387-542, 2018. [PDF]

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Figure: Design and operation of the acoustofluidic rotational manipulation (ARM) device. (a) A schematic of the experimental setup. The piezoelectric transducer that generates acoustic waves is placed adjacent to the microfluidic channel. The acoustic waves actuate air microbubbles trapped within sidewall microcavities. (b) An optical image showing a mid-L4 stage C. elegans trapped by multiple oscillating microbubbles. Scale bar, 100 microns.

The precise rotational manipulation of single cells or organisms is invaluable to many applications in biology, chemistry, physics and medicine. In this article, we describe an acoustic-based, on-chip manipulation method that can rotate single microparticles, cells and organisms. To achieve this, we trapped microbubbles within predefined sidewall microcavities inside a microchannel. In an acoustic field, trapped microbubbles were driven into oscillatory motion generating steady microvortices which were utilized to precisely rotate colloids, cells and entire organisms (that is, C. elegans). We have tested the capabilities of our method by analysing reproductive system pathologies and nervous system morphology in C. elegans. Using our device, we revealed the underlying abnormal cell fusion causing defective vulval morphology in mutant worms. Our acoustofluidic rotational manipulation (ARM) technique is an easy-to-use, compact, and biocompatible method, permitting rotation regardless of optical, magnetic or electrical properties of the sample under investigation.

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Figure: Bubble-based switching between multiple stimuli. (a) Schematic of the experimental setup for chemical switching. The microfluidic channel contains HSSs of different sizes and, subsequently, bubbles of different sizes that are independently driven by transducers bonded to the substrate adjacent to the channel. (b) Top, visualization of microstreaming from the bubble trapped in HSS A (red) while no streaming is observed in the bubble trapped in HSS B (blue) at an excitation frequency of 14.7 kHz. Bottom, visualization of the microstreaming from the bubble trapped in HSS B while no streaming occurs in HSS A at an excitation frequency of 29.5 kHz. (c) Table showing the concept of binary logic circuitry. (d) Results demonstrating switching between the blue and red dyes. (e) Graph of experimental data for switching between red and blue dyes in the selected ROI marked in panel d.

Eliciting a cellular response to a changing chemical microenvironment is central to many biological processes including gene expression, cell migration, differentiation, apoptosis, and intercellular signaling. The nature and scope of the response is highly dependent upon the spatiotemporal characteristics of the stimulus. To date, studies that investigate this phenomenon have been limited to digital (or step) chemical stimulation with little control over the temporal counterparts. Here, we demonstrate an acoustofluidic (i.e., fusion of acoustics and microfluidics) approach for generating programmable chemical waveforms that permits continuous modulation of the signal characteristics including the amplitude (i.e., sample concentration), shape, frequency, and duty cycle, with frequencies reaching up to 30 Hz. Furthermore, we show fast switching between multiple distinct stimuli, wherein the waveform of each stimulus is independently controlled. Using our device, we characterized the frequency-dependent activation and internalization of the β2-adrenergic receptor (β2-AR), a prototypic G-protein coupled receptor (GPCR), using epinephrine. The acoustofluidic-based programmable chemical waveform generation and switching method presented herein is expected to be a powerful tool for the investigation and characterization of the kinetics and other dynamic properties of many biological and biochemical processes.

References: 

1. Ahmed*, D., Ozcelik*, A., Bojanala, N., Nama, N., Upadhyay, A., Chen, Y.C., Hanna-Rose, W., and Huang, T.J., Rotational manipulation of single cells and organisms using acoustic waves, Nature Communications, Vol. 7, pp. 11085, 2016 (*equal contributions).[PDF]

2. Orbay, S., Ozcelik, A., Lata, J.P., Kaynak, M., Wu, M.X., and Huang, T.J., Mixing high-viscosity fluids via acoustically driven bubbles, Journal of Micromechanics and Microengineering, Vol. 27, pp. 015008, 2016. [PDF]

3.  Xie, Y.L., Nama, N., Li, P., Mao, Z.M., Huang, P.H., Zhao, C.L., Costanzo, F., and Huang, T.J., Probing cell deformability via acoustically actuated bubbles, Small, Vol. 12 (7), pp. 902-910, 2016.(featured as front cover image)[PDF]

4. Xie, Y.L., Ahmed, D., Lapsley, M.L., Lu, M.Q., Li, S.X., and Huang, T.J., Acoustofluidic relay: sequential trapping and transporting of microparticles via acoustically excited oscillating bubbles, Journal of Laboratory Automation, Vol. 19(2), pp. 137-143, 2014.[PDF]

5. Zhao, C.L., Xie, Y.L.,Mao, Z.M., Zhao, Y.H., Rufo, J., Yang, S.K., Guo, F., Mai, J.D., and Huang, T.J., Theory and experiment on particle trapping and manipulation via optothermally generated bubbles, Lab on a Chip, Vol. 14, pp.384-391, 2014. [PDF]

6. Ahmed, D., Muddana, H., Lu, M.Q., French, J., Ozcelik, A., Fang, Y., Butler, P., Benkovic, S., Manz, A., and Huang, T.J., An acoustofluidic chemical waveform generator and switch, Analytical Chemistry, Vol. 86, pp. 11803−11810, 2014.[PDF]

7. Xie*, Y.L., Zhao*, C.L., Zhao, Y.H., Li, S.X., Rufo, J., Yang, S.K., Guo, F., and Huang, T.J., Optoacoustic tweezers: a programmable, localized cell concentrator based on opto-thermally generated, acoustically activated, surface bubbles, Lab on a Chip, Vol. 13, pp. 1772-1779, 2013.(*equal contributions)(featured as front cover image)[PDF]

8. Ahmed, D., Chan,C.Y., Lin, S.S., Muddana, H.S., Nama, N., Benkovic, S.J. and Huang, T.J., Tunable, Pulsatile Chemical Gradient Generation via Acoustically Driven Oscillating Bubbles, Lab on a Chip, Vol. 13, pp. 328-331, 2013.(featured as front cover image)[PDF]

9. Huang, P.H., Lapsley, M.I., Ahmed, D., Chen, Y.C., Wang, L., and Huang, T.J., A Single-Layer, Planar, Optofluidic Switch Powered By Acoustically Driven, Oscillating Microbubbles, Applied Physics Letters, Vol.101, 141101, 2012. (featured as front cover image) [PDF]

10. Ahmed, D., Mao, X.L., Juluri, B.K. and Huang, T.J., A Fast Microfluidic Mixer Based on Acoustically Driven Sidewall-Trapped Microbubbles, Microfluidics and Nanofluidics, Vol. 7, pp. 727-731, 2009. [PDF]

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Figure: (a) Schematic of the sharp-edge-based acoustofluidic mixing device. This device includes a PDMS microfluidic channel and a piezoelectric transducer. (b) Schematic showing the acoustic streaming phenomenon around the tip of an acoustically oscillated sharp-edge. (c) Schematic showing the design of the channel and sharp-edge.

Rapid and homogeneous mixing inside a microfluidic channel is demonstrated via the acoustic streaming phenomenon induced by the oscillation of sidewall sharp-edges. By optimizing the design of the sharpedges, excellent mixing performance and fast mixing speed can be achieved in a simple device, making our sharp-edge-based acoustic micromixer a promising candidate for a wide variety of applications.

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Figure: (a) Schematic of the sharp-edge-based acoustofluidic pumping device. This device includes a PDMS microfluidic channel and a piezoelectric transducer. (b) Schematic showing the acoustic streaming phenomenon around the tip of a tilted oscillating sharp-edge structure. (c) Schematic showing the design of the channel and sharp-edge structure.

We present a programmable acoustofluidic pump that utilizes the acoustic streaming effects generated by the oscillation of tilted sharp-edge structures. This sharp-edge-based acoustofluidic pump is capable of generating stable flow rates as high as 8 μL min−1 (~76 Pa of pumping pressure), and it can tune flow rates across a wide range (nanoliters to microliters per minute). Along with its ability to reliably produce stable and tunable flow rates, the acoustofluidic pump is easy to operate and requires minimum hardware, showing great potential for a variety of applications.

References:

1. Nama, N., Huang, P.H., Huang, T.J., and Costanzo, F., Investigation of micromixing by acoustically oscillated sharp-edges, Biomicrofluidics, Vol. 10, 024124, 2016.[PDF]

2. Ozcelik, A., Nama, N., Huang, P.H., Kaynak, M., McReynolds, M.R., Hanna‐Rose, W., and Huang, T.J., Acoustofluidic rotational manipulation using oscillating solid structures, Small, Vol. 12, pp. 5120-5125, 2016.(featured as front cover image) [PDF]

3. Huang, P.H., Chan, C.Y., Li, P., Nama, N., Xie, Y.L.,  Wei, C.H., Chen, Y.C., Ahmed, D., and Huang, T.J., A spatiotemporally controllable chemical gradient generator via acoustically oscillating sharp-edge structures, Lab on a Chip, Vol. 15, pp. 4166-4176, 2015.[PDF]

4. Huang, P.H., Ren, L.Q., Nama, N., Li, S.X.,Li, P., Yao, X.G., Cuento, R.A., Wei, C.H., Chen, Y.C., Xie, Y.L., Nawaz, A.A., Alevy, Y.G., Holtzman, M.J., McCoy, J.P., Levine, S.J., and Huang, T.J., An acoustofluidic sputum liquefier, Lab on a Chip, Vol. 15, pp. 3125-3131, 2015.(featured as front cover image)[PDF]

5. Nama, N., Huang, P.H., Huang, T.J., and Costanzo, F., Investigation of acoustic streaming patterns around oscillating sharp edges, Lab on a Chip, Vol. 14, pp. 2824-2836, 2014.[PDF]

6. Huang, P.H., Nama, N., Mao, Z.M., Li, P., Rufo, J., Chen, Y.C., Xie, Y.L., Wei, C.H., Wang, L., and Huang, T.J., A reliable, programmable acoustofluidic pump powered by oscillating sharp-edge structures, Lab on a Chip, Vol. 14, pp. 4319-4323, 2014. [PDF]

7. Huang, P.H., Xie, Y.L., Ahmed, D., Rufo, J., Nama, N., Chen, Y.C., Chan, C.Y., and Huang, T.J., An acoustofluidic micromixer based on oscillating sidewall sharp-edges, Lab on a Chip, Vol. 13, pp. 3847-3852, 2013.[PDF]

Harmonic acoustics for dynamic and selective particle manipulation

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Figure: a, Schematic of HANDS manipulation with Fourier-synthesized harmonic acoustic waves. b, Top, a colloidal crystal with controllable particle numbers and conformation can be assembled. Bottom, large area patterning with a tuneable lattice constant can be generated for colloidal clusters or single cells. c, The modulation of frequency and amplitude of harmonic waves can generate soft lattices and reconfigurable colloidal crystal or tuneable patterning of single particles or cells. d, Fluorescent imaging showing selective pairing of two U937 cells among six cells by localized modulation of the intercellular distance. The positions of the cells are indicated by the fluorescent intensity profile (averaged from five cell groups). Scale bars, 10 μm. e, Comparison of two patterned colloidal clusters with equal trapping spacing and tuneable trapping spacings, as analytically simulated and experimentally generated using HANDS manipulation. Colloidal clusters are formed with 2 μm fluorescent polystyrene particles in each trapping acoustic wells. Scale bars, 20 μm.

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.

Acousto-dielectric tweezers enable independent manipulation of multiple particles

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Figure: Schematic illustrating the mechanism of acousto-dielectric tweezers.

Acoustic tweezers have gained substantial interest in biology, engineering, and materials science for their label-free, precise, contactless, and programmable manipulation of small objects. However, acoustic tweezers cannot independently manipulate multiple microparticles simultaneously. This study introduces acousto-dielectric tweezers capable of independently manipulating multiple microparticles and precise control over intercellular distances and cyclical cell pairing and separation for detailed cell-cell interaction analysis. Our acousto-dielectric tweezers leverage the competition between acoustic radiation forces, generated by standing surface acoustic waves (SAWs), and dielectrophoretic (DEP) forces, induced by gradient electric fields. Modulating these fields allows for the precise positioning of individual microparticles at points where acoustic radiation and DEP forces are in equilibrium. This mechanism enables the simultaneous movement of multiple microparticles along specified paths as well as cyclical cell pairing and separation. We anticipate our acousto-dielectric tweezers to have enormous potential in colloidal assembly, cell-cell interaction studies, disease diagnostics, and tissue engineering.

Acoustic tweezers: patterning cells and microparticles using standing surface acoustic waves (SSAW)

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Figure: Patterning of fluorescent polystyrene microbeads. Optical images of the ‘‘acoustic tweezers’’ devices used in (a) 1D and (b) 2D patterning experiments, respectively. (c) Distribution of the microbeads before and after the 1D patterning process. The microchannel (width 150 μm and depth 80 μm) covered three lines of pressure nodes of the generated SSAW. The wavelength of SAW was 100 mm (d), Distribution of the microbeads before and after the 2D patterning process. The SAW wavelength was 200 μm.

Here we present an active patterning technique named ‘‘acoustic tweezers’’ that utilizes standing surface acoustic wave (SSAW) to manipulate and pattern cells and microparticles. This technique is capable of patterning cells and microparticles regardless of shape, size, charge or polarity. Its power intensity, approximately 5 x 105 times lower than that of optical tweezers, compares favorably with those of other active patterning methods. Flow cytometry studies have revealed it to be non-invasive. The aforementioned advantages, along with this technique’s simple design and ability to be miniaturized, render the ‘‘acoustic tweezers’’ technique a promising tool for various applications in biology, chemistry, engineering, and materials science.

On-Chip Manipulation of Single Microparticles, Cells, and Organisms Using Surface Acoustic Waves

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Figure: Device structure and working mechanism of the acoustic tweezers. (A) Schematic illustrating a microfluidic device with orthogonal pairs of chirped IDTs for generating standing SAW. (B) A standing SAW field generated by driving chirped IDTs at frequency f 1 and f2. When particles are trapped at the nth pressure node, they can be translated a distance of (Δλ∕2) n by switching from f1 to f2. This relationship indicates that the particle displacement can be tuned by varying the pressure node where the particle is trapped.

Techniques that can dexterously manipulate single particles, cells, and organisms are invaluable for many applications in biology, chemistry, engineering, and physics. Here, we demonstrate standing surface acoustic wave based “acoustic tweezers” that can trap and manipulate single microparticles, cells, and entire organisms (i.e., Caenorhabditis elegans) in a single-layer microfluidic chip. Our acoustic tweezers utilize the wide resonance band of chirped interdigital transducers to achieve real-time control of a standing surface acoustic wave field, which enables flexible manipulation of most known microparticles. The power density required by our acoustic device is significantly lower than its optical counterparts (10,000,000 times less than optical tweezers and 100 times less than optoelectronic tweezers), which renders the technique more biocompatible and amenable to miniaturization. Cell-viability tests were conducted to verify the tweezers’ compatibility with biological objects. With its advantages in biocompatibility, miniaturization, and versatility, the acoustic tweezers presented here will become a powerful tool for many disciplines of science and engineering.

Standing surface acoustic wave (SSAW)-based microfluidic cytometer

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Figure: (A) SSAW-based 3D particle focusing observed from different perspectives (I: top view, II: side view). (B) Top-view bright field images and (C) side-view fluorescent stacking images of the particle flow pattern without (OFF) and with (ON) SSAW focusing. The dashed lines in (C) indicate the channel boundaries. (D) Top-view images showing cell focusing by SSAW.

The development of microfluidic chip-based cytometers has become an important area due to their advantages of compact size and low cost. Herein, we demonstrate a sheathless microfluidic cytometer which integrates a standing surface acoustic wave (SSAW)-based microdevice capable of 3D particle/cell focusing with a laser-induced fluorescence (LIF) detection system. Using SSAW, our microfluidic cytometer was able to continuously focus microparticles/cells at the pressure node inside a microchannel. Flow cytometry was successfully demonstrated using this system with a coefficient of variation (CV) of less than 10% at a throughput of ~1000 events s−1 when calibration beads were used. We also demonstrated that fluorescently labeled human promyelocytic leukemia cells (HL-60) could be effectively focused and detected with our SSAW-based system. This SSAW-based microfluidic cytometer did not require any sheath flows or complex structures, and it allowed for simple operation over a wide range of sample flow rates. Moreover, with the gentle, bio-compatible nature of low-power surface acoustic waves, this technique is expected to be able to preserve the integrity of cells and other bioparticles.

Cell Separation Using Tilted-Angle Standing Surface Acoustic Waves

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Figure: Schematic illustration of working principle and device structure. (A) Photo showing a taSSAW-based cell-separation device. (B) and (C) Separation process for 10- and 2-μm-diameter polystyrene beads in the taSSAW working region and the outlet region, respectively.

We have developed a unique approach for the separation of particles and biological cells through standing surface acoustic waves oriented at an optimum angle to the fluid flow direction in a microfluidic device. This experimental setup, optimized by systematic analyses, has been used to demonstrate effective separation based on size, compressibility, and mechanical properties of particles and cells. The potential of this method for biological–biomedical applications was demonstrated through the example of isolating MCF-7 breast cancer cells from white blood cells. The method offers a possible route for label-free particle or cell separation for many applications in research, disease diagnosis, and drug-efficacy assessment.

A high-throughput acoustic cell sorter

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Figure: (A) Schematic of the SSAW-based sorter excited by FIDTs. (B) The concentric geometry of the FIDTs. θ is the degree of arcs and R is the radius of innermost transducer. O is the focal point and the two sets of FIDTs is O-axis symmetric. (C) An optical image of our high throughput SSAW sorter.

Acoustic-based fluorescence activated cell sorters (FACS) have drawn increased attention in recent years due to their versatility, high biocompatibility, high controllability, and simple design. However, the sorting throughput for existing acoustic cell sorters is far from optimum for practical applications. Here we report a high-throughput cell sorting method based on standing surface acoustic waves (SSAWs). We utilized a pair of focused interdigital transducers (FIDTs) to generate SSAW with high resolution and high energy efficiency. As a result, the sorting throughput is improved significantly from conventional acoustic-based cell sorting methods. We demonstrated the successful sorting of 10 μm polystyrene particles with a minimum actuation time of 72 μs, which translates to a potential sorting rate of more than 13,800 events per second. Without using a cell-detection unit, we were able to demonstrate an actual sorting throughput of 3,300 events per second. Our sorting method can be conveniently integrated with upstream detection units, and it represents an important development towards a functional acoustic-based FACS system.

Controlling Cell-Cell Interactions using Surface Acoustic Waves

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Figure: Translation of suspended assemblies to the adherent state. (A) Schematic of experimental setup and procedure. Linear assemblies of cells were formed under the control of a tunable acoustic well. After the removal of the acoustic field, cells were allowed to drop to the surface and attach. (B) The process of in-suspension assembly and attachment. HEK 293T cells were first assembled in suspension. The acoustic field was maintained for 1 h in order that cells could be kept in suspension regardless of the properties of the surface. Once the acoustic field is removed, cells quickly attach to the collagen-coated surface and start to spread while maintaining the same assembly as in suspension. In-suspension assembly and attachment of (C) HMVEC and (E) HeLa S3 cells, respectively. Dye transfer between attached (D) HMVEC and (F) HeLa S3 cells, respectively, with defined geometry. Scale bar: 20 μm.

We present a unique acoustic well approach that can precisely control cell-to-cell distance and cell–cell interactions. Our technology can achieve high precision and high throughput simultaneously while preserving the integrity of cells. It is capable of creating cell assemblies with precise spatial control both in suspension and on a substrate. We envision the exploitation of this powerful technology, for example, in the study of cell–cell interactions in fields, such as immunology, developmental biology, neuroscience, and cancer metastasis, and in the studies of cell–cell and cell–matrix adhesion.

Acoustic Separation of Circulating Tumor Cells

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Figure: Schematic illustration and image of the high-throughput taSSAW device for cancer cell separation. (A) Illustration of taSSAW-based cell separation. (B) Schematic of the working mechanism behind taSSAW-based cell separation. The direction of the pressure nodes and pressure antinodes were established at an angle of inclination (θ) to the fluid flow direction inside a microfluidic channel. Larger CTCs experience a larger acoustic radiation force (Fac) than WBCs (Faw). As a result, CTCs have a larger vertical displacement (normal to the flow direction) than WBCs. Fdc and Fdw are the drag force experienced by CTCs and WBCs, respectively. (C) An actual image of the taSSAW cell separation device. Blue ink was used to help visualize the microfluidic channel.

The separation and analysis of circulating tumor cells (CTCs) provides physicians a minimally invasive way to monitor the response of cancer patients to various treatments. Among the existing cellseparation methods, acoustic-based approaches provide significant potential to preserve the phenotypic and genotypic characteristics of sorted cells, owing to their safe, label-free, and contactless nature. In this work, we report the development of an acoustic-based device that successfully demonstrates the isolation of rare CTCs from the clinical blood samples of cancer patients. Our work thus provides a unique means to obtain viable and undamaged CTCs, which can subsequently be cultured. The results presented here offer unique pathways for better cancer diagnosis, prognosis, therapy monitoring, and metastasis research.

Three-dimensional manipulation of single cells using surface acoustic waves

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Figure: Illustration of our 3D acoustic tweezers. (A) Configuration of the planar surface acoustic wave generators, used to generate volumetric nodes, surrounding the microfluidic experimental area. The Inset indicates a single particle within a “3D trapping node,” which is independently manipulated along the x, y, or z axes. (B) Numerical simulation results mapping the acoustic field around a particle that shows the physical operating principle for the 3D acoustic tweezers. The 3D trapping node in the microfluidic chamber is created by two superimposed, orthogonal, standing surface acoustic waves and the induced acoustic streaming.

We present 3D acoustic tweezers, which can trap and manipulate single cells and particles along three mutually orthogonal axes of motion by recourse to surface acoustic waves. We use 3D acoustic tweezers to pick up single cells, or entire cell assemblies, and deliver them to desired locations to create 2D and 3D cell patterns, or print the cells into complex shapes. This technology is thus shown to offer better performance over prior cell manipulation techniques in terms of both accurate and precise motion in a noninvasive, label-free, and contactless manner. This method offers the potential to accurately print 3D multicellular architectures for applications in biomanufacturing, tissue engineering, regenerative medicine, neuroscience, and cancer metastasis research.

References: 

1. Barnkob, R., Nama, N., Ren, L.Q., Huang, T.J., Costanzo, F., and Kahler, C.J., Acoustically Driven Fluid and Particle Motion in Confined and Leaky Systems, Physical Review Applied, Vol. 9, pp. 014027, 2018. [PDF]

2. Ren, L.Q., Zhou, D.K., Mao, Z.M., Xu, P.T., Huang, T.J., and Mallouk, T.E., Rheotaxis of Bimetallic Micromotors Driven by Chemical−Acoustic Hybrid Powervirology, ACS Nano, Vol. 11, pp. 10691-10698, 2017. [PDF]

3. Wu, M.X., Ouyang, Y.S., Wang, Z.Y., Zhang, R., Huang, P.H., Chen, C.Y., Li, H., Li, P., Quinn, D., Dao, M., Suresh, S., Sadovsky, Y., and Huang, T.J.,  Isolation of exosomes from whole blood by integrating acoustics and microfluidics, Proceedings of the National Academy of Sciences of the United States of America (PNAS), Vol. 114, pp. 10584-10589, 2017. [PDF] (See video news from MIT, Duke, and Science channel)

4. Wu, M.X., Mao, Z.M., Chen, K.J., Bachman, H., Chen, Y.C., Rufo, J., Ren, L.Q., Li, P., Wang, L., and Huang, T.J., Acoustic Separation of Nanoparticles in Continuous Flow, Advanced Functional Materials, Vol. 27, pp. 1606039, 2017. (featured as back cover image) [PDF] 

5. Lata, J.P., Guo, F., Guo, J.S., Huang, P.H., Yang, J., and Huang, T.J., Surface acoustic waves grant superior spatial control of cells embedded in hydrogel fibers, Advanced Materials, Vol. 28, pp. 8632-8638, 2016.(featured as front cover image) [PDF] 

6. Chen Y.C., Wu, M.X., Ren, L.Q., Liu, J.Y., Whitley, P.H., Wang, L., and Huang, T.J., High-throughput acoustic separation of platelets from whole blood, Lab on a Chip, Vol. 16, pp. 3466-3472, 2016. [PDF]

7. Chen, K.J., Wu, M.X., Guo, F., Li, P., Chan, C.Y., Mao, Z.M. Li, S.X., Ren, L.Q., Zhang, R., and Huang, T.J., Rapid formation of size-controllable multicellular spheroids via 3D acoustic tweezers, Lab on a Chip, Vol. 16, pp. 2636-2643, 2016.[PDF]

8. Guo, F., Mao, Z.M., Chen, Y.C., Xie, Z.W., Lata, J.P., Li, P., Ren, L.Q., Liu, J.Y., Yang, J., Daoc, M., Sureshd, S., and Huang, T.J., Three-dimensional Manipulation of Single Cells Using Surface Acoustic Waves, Proceedings of the National Academy of Sciences of the United States of America (PNAS), Vol. 113, pp. 1522-1527, 2016.[PDF](“Highly Cited Paper” by Thomson Reuters—top 1% of highest cited papers in Chemistry)

9. Mao, Z.M., Xie, Y.L., Guo, F., Ren, L.Q., Huang, P.H., Chen, Y.C., Rufo, J., Costanzo, F., and Huang, T.J., Experimental and numerical studies on standing surface acoustic wave microfluidics, Lab on a Chip, Vol. 16, pp. 515-524, 2016.[PDF]

10. Guo, F., Xie. Y.L., Li, S.X., Lata, J., Ren, L.Q.,Mao, Z.M., Ren, B.Y., Wu, M.X., Ozcelik, A., and Huang, T.J., Reusable acoustic tweezers for disposable devices, Lab on a Chip, Vol. 15, pp. 4517 - 4523, 2015.(featured as front cover image)[PDF]

11. Ren, L.Q., Chen, Y.C., Li, P., Mao, Z.M., Huang, P.H., Rufo, J., Guo, F., Wang, L.,McCoy, J.P.,  Levine, S.J., and Huang, T.J., A high-throughput acoustic cell sorter, Lab on a Chip, Vol. 15, pp. 3870-3879, 2015.(featured as front cover image)[PDF]

12. Guo, F., Zhou, W.J., Li, P.,  Mao, Z.M., Yennawar, N., French, J.B., and Huang, T.J., Precise Manipulation and Patterning of Protein Crystals for Macromolecular Crystallography using Surface Acoustic Waves, Small, Vol. 11 (23), pp. 2733–2737, 2015.(featured as front cover image)[PDF]

13. Nama, N.,Barnkob, R., Mao, Z.M., Kähler, C.J., Costanzo, F., and Huang, T.J., Numerical study of acoustophoretic motion of particles in a PDMS microchannel driven by surface acoustic waves, Lab on a Chip, Vol. 15, pp. 2700-2709, 2015.[PDF]

14. Li, P., Mao, Z.M., Peng, Z.L., Zhou, L.L., Chen, Y.C., Huang, P.H., Truic, C.I., Drabick, J.J., El-Deiry, W.S., Dao, M., Suresh, S., and Huang, T.J., Acoustic separation of circulating tumor cells, Proceedings of the National Academy of Sciences of the United States of America (PNAS), Vol. 112, pp. 4970-4975, 2015.[PDF](“Highly Cited Paper” by Thomson Reuters—top 1% of highest cited papers in Chemistry)

15. Guo, F., Li, P.,  French, J.B., Mao, Z.M., Zhao, H., Li, S.X., Nama, N., Fick, J.R., Benkovic, S.J., and Huang, T.J., Controlling Cell-Cell Interactions using Surface Acoustic Waves, Proceedings of the National Academy of Sciences of the United States of America (PNAS), Vol. 112, pp. 43–48, 2015.[PDF](“Highly Cited Paper” by Thomson Reuters—top 1% of highest cited papers in Molecular Biology & Genetics)

16. Li, S.X., Ding, X.Y., Mao, Z.M., Chen, Y.C., Nama, N., Guo, F., Li, P., Wang, L., Cameron, C.E. and Huang, T.J., Standing surface acoustic wave (SSAW)-based cell washing, Lab on a Chip, Vol. 15, pp. 331-338, 2015.[PDF]

17. Li, S.X., Guo, F., Chen, Y.C., Ding, X.Y., Li, P., Wang, L., Cameron, C.E., and Huang, T.J., Standing surface acoustic wave (SSAW)-based cell co-culture, Analytical Chemistry, Vol. 86 (19), pp 9853–9859, 2014.[PDF]

18. Ding, X.Y., Peng, Z.L., Lin, S.S., Geri, M., Li, S.X., Li, S.X., Chen, Y.C., Dao, M., Suresh, S., and Huang, T.J., Cell separation using tilted-angle standing surface acoustic waves, Proceedings of the National Academy of Sciences of the United States of America (PNAS), Vol. 111, pp. 12992-12997, 2014.[PDF](“Highly Cited Paper” by Thomson Reuters—top 1% of highest cited papers in Chemistry)

19. Chen, Y.C.,Li, S.X., Gu, Y.Y., Li, P., Ding, X.Y., McCoy, J.P., Levine, S.J., Wang, L., and Huang, T.J., Continuous enrichment of low-abundance cell sample using standing surface acoustic waves (SSAW), Lab on a Chip, Vol. 14, pp. 924-930, 2014.[PDF]

20. Chen, Y.C., Nawaz, A.A., Zhao, Y.H., Huang, P.H., McCoy, J.P., Levine, S.J., Wang, L., and Huang, T.J., Standing surface acoustic wave (SSAW)-based microfluidic cytometer, Lab on a Chip, Vol. 14, pp. 916-923, 2014.[PDF]

21. Ding, X.Y., Li, P., Lin, S.S., Stratton, Z.S., Nama, N., Guo, F., Slotcavage,D.,  Mao, X.L., Shi, J.J., Costanzo, F., and Huang, T.J.,Surface acoustic wave microfluidics, Lab on a Chip, Vol. 13, pp. 3626-3649, 2013.[PDF](“Highly Cited Paper” by Thomson Reuters—top 1% of highest cited papers in Chemistry)

22. Liu, Y.J., Lu, M.Q., Ding, X.Y., Leong, E.S.P., Lin, S.S., Shi, J.J., Teng, J.H., Wang, L., Bunning, T.J., and Huang, T.J.,Holographically Formed, Acoustically Switchable Gratings Based on Polymer-Dispersed Liquid Crystals, Journal of Laboratory Automation, Vol. 18, pp. 291-295, 2013.(featured as front cover image)[PDF]

23. Li, S.X., Ding, X.Y., Guo, F., Chen, Y.C., Lapsley, M., Lin, S.S., Wang, L., McCoy, J.P., Cameron,C., and Huang, T.J., An on-chip, multichannel droplet sorter using standing surface acoustic waves (SSAW), Analytical Chemistry, Vol. 85, pp. 5468-5474, 2013.[PDF]

24. Chen, Y.C., Ding, X.Y., Lin, S.S., Yang, S.S., Huang, P.H., Nama, H., Zhao, Y.H., Nawaz, A.A., Guo ,F., Wang ,W., Gu, Y.Y., Mallouk, T.E., and Huang, T.J., Tunable Nanowire Patterning Using Standing Surface Acoustic Waves, ACS Nano, Vol. 7, pp. 3306-3314, 2013.[PDF]

25. Ding, X.Y.,  Lin, S.S., Kiraly, B., Yue, H.J., Li, S.X., Shi, J.J., Benkovic, S.J., and Huang, T.J., On-Chip Manipulation of Single Microparticles, Cells, and Organisms Using Surface Acoustic Waves, Proceedings of the National Academy of Sciences of the United States of America (PNAS), Vol. 109, pp. 11105-11109, 2012. [PDF](“Highly Cited Paper” by Thomson Reuters—top 1% of highest cited papers in Chemistry)

26. Ding, X.Y., Lin, S.S., Lapsley, M.I., Li, S.X., Guo, X., Chan, C.Y.K., Chiang, I.K., McCoy, J.P., and Huang, T.J., Standing Surface Acoustic Wave (SSAW) Based Multichannel Cell Sorting, Lab on a Chip, Vol.12, pp. 4228–4231, 2012. (featured as front cover image) [PDF]

27. Lin, S.S., Mao, X.L., and Huang, T.J., Surface Acoustic Wave (SAW) Acoustophoresis: Now and Beyond, Lab on a Chip, Vol.12, pp. 2766-2770, 2012. [PDF]

28. Ding, X.Y., Shi, J.J., Lin, S.S., Yazdi, S., Kiraly, B., and Huang, T.J., Tunable Patterning of Microparticles and Cells using Standing Surface Acoustic Waves, Lab on a Chip, Vol. 12, pp. 2491-2497, 2012. (featured as back cover image) [PDF]

29. Shi, J.J., Yazdi, S., Lin,S.S., Ding, X.Y., Chiang, I.K., Sharp, K., and Huang, T.J., Three-Dimensional Continuous Particle Focusing in a Microfluidic Channel via Standing Surface Acoustic Waves (SSAW), Lab on a Chip, Vol. 11, pp. 2319-2324, 2011. (featured as front cover image) [PDF]

30. Liu, Y.J., Ding, X.Y., Lin, S.S., Shi, J.J., Chiang, I.K., and Huang, T.J., Surface Acoustic Wave Driven Light Shutters Using Polymer-Dispersed Liquid Crystals, Advanced Materials, Vol. 23, pp. 1656-1659, 2011. (featured as front cover image) [PDF]

31. Shi, J.J., Huang, H., Stratton, Z., Lawit, A., Huang, Y.P. and Huang, T.J., Continuous Particle Separation in a Microfluidic Channel via Standing Surface Acoustic Waves (SSAW), Lab on a Chip, Vol. 9, pp. 3354-3359, 2009. (featured as back cover image) [PDF](“Highly Cited Paper” by Thomson Reuters—top 1% of highest cited papers in Chemistry)

32. Shi, J.J., Ahmed, D., Mao, X.L., Lin, S.S., and Huang, T. J., Acoustic Tweezers: Patterning Cells and Microparticles Using Standing Surface Acoustic Waves (SSAW), Lab on a Chip, Vol. 9, pp. 2890-2895, 2009. (featured as front cover image) [PDF](“Highly Cited Paper” by Thomson Reuters—top 1% of highest cited papers in Chemistry)

33. Shi, J.J., Mao, X.L., Ahmed, D., Colletti, A., Huang, T.J., Focusing Microparticles in a Microfluidic Channel with Standing Surface Acoustic Waves (SSAW), Lab on a Chip, Vol. 8, pp. 221-223, 2008. [PDF](“Highly Cited Paper” by Thomson Reuters—top 1% of highest cited papers in Chemistry)

Introduction:

Optofluidics refers to manipulation of light using fluids, or vice-verse, on the micro to nano meter scale. By taking advantage of the microfluidic manipulation, the optical properties of the fluids can be precisely and flexibly controlled to realize reconfigurable optical components which are otherwise difficult or impossible to implement with solid-state technology. In addition, the unique behavior of fluids on micro/nano scale has given rise to the possibility to manipulate the fluid using light.

Due to their unique electronic, magnetic, and optical properties compared with bulk materials, nanomaterials and nanostructures have tremendous potential in many applications. We are devoted to the development of novel nanostructures, which offer interesting photonic properties and could be readily applied to medical diagnostics and therapeutics. These nanostructures will bridge the interface between modern molecular biology and nanotechnology.

In the Duke Acoustofluidcis laboratory, we aim to (1) investigate interactions of light and metal at nanoscale to address fundamental issues of nanophotonics, (2) develop cost-effective and high-throughput nanofabrication techniques and nanoengineering methods to produce plasmonic nanostructures for different applications, and (3) design, model and fabricate active plasmonic devices to benefit information technology, and medical diagnosis and therapy.

Below are several examples related to optofluidics and plasmofluidics:

A reconfigurable plasmofluidic lens

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Figure: Schematic of the reconfigurable plasmofluidic lens, wherein a laser-induced surface bubble is used to control the propagation of SPPs at the metal surface. (c–e) Surface bubbles with different diameters (18, 14 and 6 microns, respectively) generated on the gold film. The white dashed circle represents the surface bubble boundary on gold film. Scale bar, 10 microns.

Plasmonics provides an unparalleled method for manipulating light beyond the diffraction limit, making it a promising technology for the development of ultra-small, ultra-fast and power-efficient optical devices. To date, the majority of plasmonic devices are in the solid state and have limited tunability or configurability. Moreover, individual solid-state plasmonic devices lack the ability to deliver multiple functionalities. Here we utilize laser-induced surface bubbles on a metal film to demonstrate, for the first time, a plasmonic lens in a microfluidic environment. Our ‘plasmofluidic lens’ is dynamically tunable and reconfigurable. We record divergence, collimation and focusing of surface plasmon polaritons using this device. The plasmofluidic lens requires no sophisticated nanofabrication and utilizes only a single low-cost diode laser. Our results show that the integration of plasmonics and micro- fluidics allows for new opportunities in developing complex plasmonic elements with multiple functionalities, high-sensitivity and high-throughput biomedical detection systems, as well as on-chip, all-optical information processing techniques.

Tunable optofluidic cylindrical microlens

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Figure: The 3D architectures of water flow obtained with confocal microscopy at the flow rates of (a) 100 ml min21 , (b) 150 ml min21 , and (c) 250 ml min21 . Images in (d–f) show the corresponding CFDsimulated 3D fluidic interfaces (isosurface of CaCl2 concentration = 2.5 M).

We report the design, fabrication, and characterization of a tunable optofluidic microlens that focuses light within a microfluidic device. The microlens is generated by the interface of two co-injected miscible fluids of different refractive indices, a 5 M CaCl2 solution (nD = 1.445) and deionized (DI) water (nD = 1.335). When the liquids flow through a 90-degree curve in a microchannel, a centrifugal effect causes the fluidic interface to be distorted and the CaCl2 solution bows outwards into the DI water portion. The bowed fluidic interface, coupled with the refractive index contrast between the two fluids, yields a reliable cylindrical microlens. The optical characteristics of the microlens are governed by the shape of the fluidic interface, which can be altered by simply changing the flow rate. Higher flow rates generate a microlens with larger curvature and hence shorter focal length. The changing of microlens profile is studied using both computational fluid dynamics (CFD) and confocal microscopy. The focusing effect is experimentally characterized through intensity measurements and image analysis of the focused light beam, and the experimental data are further confirmed by the results from a ray-tracing optical simulation. Our investigation reveals a simple, robust, and effective mechanism for integrating optofluidic tunable microlenses in lab-on-a-chip systems.

Hydrodynamically Tunable Optofluidic Cylindrical Microlens

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Figure: Principle and design of the L-GRIN lens. (A) A schematic diagram showing the comparison between the classic refractive lens (A1) and GRIN lens (A2). Change of the refractive index contrast in GRIN lens can result in change of focal distance (A2–A3), and shift of optical axis can result in change of output light direction (A4). (B) Schematic of the L-GRIN lens design (B1), microscopic image of the L-GRIN lens in operation (B2, left), and the expected refractive index distribution at two locations (I and II) inside the lens (B2, right). High optical contrast areas (dark streaks) were observed near the fluidic boundaries (B2, left), suggesting significant variation of refractive index due to the CaCl2 diffusion. (C) Schematic drawing showing two operation modes of the L-GRIN lens: the translation mode with variable focal length including no-focusing (C1), a large focal distance (C2), and a small focal distance (C3); and the swing mode with variable output light direction (C3–C5).

We report a tunable optofluidic microlens configuration named the Liquid Gradient Refractive Index (L-GRIN) lens for focusing light within a microfluidic device. The focusing of light was achieved through the gradient refractive index (GRIN) within the liquid medium, rather than via curved refractive lens surfaces. The diffusion of solute (CaCl2) between side-by-side co-injected microfluidic laminar flows was utilized to establish a hyperbolic secant (HS) refractive index profile to focus light. Tailoring the refractive index profile by adjusting the flow conditions enables not only tuning of the focal distance (translation mode), but also shifting of the output light direction (swing mode), a second degree of freedom that to our knowledge has yet to be accomplished for in-plane tunable microlenses. Advantages of the L-GRIN lens also include a low fluid consumption rate, competitive focusing performance, and high compatibility with existing microfluidic devices. This work provides a new strategy for developing integrative tunable microlenses for a variety of lab-on-a-chip applications.

References: 

1. Mao, Z.M., Guo, F., Xie, Y.L., Zhao, Y.H., Lapsley, M.I., Wang, L., Mai, J.D., Costanzo, F., and Huang, T.J., Label-free measurements of reaction kinetics using a droplet-based optofluidic device, Journal of Laboratory Automation, Vol. 20, pp. 17-24, 2015.(featured as front cover image)[PDF]

2. Zhao, C.L., Xie, Y.L.,Mao, Z.M., Zhao, Y.H., Rufo, J., Yang, S.K., Guo, F., Mai, J.D., and Huang, T.J., Theory and experiment on particle trapping and manipulation via optothermally generated bubbles, Lab on a Chip, Vol. 14, pp.384-391, 2014.[PDF]

3. Xie*, Y.L., Zhao*, C.L., Zhao, Y.H., Li, S.X., Rufo, J., Yang, S.K., Guo, F., and Huang, T.J., Optoacoustic tweezers: a programmable, localized cell concentrator based on opto-thermally generated, acoustically activated, surface bubbles, Lab on a Chip, Vol. 13, pp. 1772-1779, 2013.(*equal contributions)(featured as front cover image)[PDF]

4. Zhao, Y.H., Stratton, Z.S., Guo, F., Lapsley, M.I., Chan, C.Y., Lin, S.S., and Huang, T.J., Optofluidic Imaging: Now and Beyond, Lab on a Chip, Vol. 13, pp. 17-24, 2013.[PDF]

5. Guo, F., Lapsley, M.I., Nawaz, A.A., Zhao, Y.H., Lin, S.S., Chen, Y.C., Yang, S.C., Zhao, X.Z., and Huang, T.J., A Droplet-based, Optofluidic Device for High-throughput, Quantitative Bioanalysis, Analytical Chemistry,Vol. 84, pp. 10745−10749, 2012.[PDF]

6. Huang, P.H., Lapsley, M.I., Ahmed, D., Chen, Y.C., Wang, L., and Huang, T.J., A Single-Layer, Planar, Optofluidic Switch Powered By Acoustically Driven, Oscillating Microbubbles, Applied Physics Letters, Vol.101, 141101, 2012. (featured as front cover image) [PDF]

7. Lapsley, M.I., Chiang, I.K., Zheng, Y.B., Ding, X.Y., Mao, X.L., and Huang, T.J., A Single-Layer, Planar, Optofluidic Mach-Zehnder Interferometer for Label-Free Detection, Lab on a Chip, Vol. 11, pp. 1795–1800, 2011.[PDF]

8. Mao, X.L., Stratton, Z.I., Nawaz, A.A., Lin, S.S., and Huang, T.J., Optofluidic Tunable Microlens by Manipulating the Liquid Meniscus Using a Flared Microfluidic Structure, Biomicrofluidics, Vol. 4, pp. 043007, 2010. (featured as front cover image) [PDF]

9. Shi, J.J., Stratton, Z., Lin, S.S., Huang, H. and Huang, T.J., Tunable Optofluidic Microlens through Active Pressure Control of an Air-Liquid Interface,Microfluidics and Nanofluidics, Vol. 9, pp. 313-318, 2010. [PDF]

10. Mao, X.L., Waldeisen, J.R., Juluri, B.K., Huang, T.J., Hydrodynamically Tunable Optofluidic Cylindrical Microlens, Lab on a Chip, Vol. 7, pp. 1303-1308, 2007. [PDF]

11. Zhang, B., Bian, Y.S., Ren, L.Q., Guo, F., Tang, S.Y., Mao, Z.M., Liu, X.M., Sun, J.J., Gong, J.Y., Guo, X.S., and Huang, T.J., Hybrid Dielectric-loaded Nanoridge Plasmonic Waveguide for Low-Loss Light Transmission at the Subwavelength Scale, Scientific Reports, Vol. 7, pp. 40479, 2017. [PDF]

12. Ahmed, D., Peng, X.L., Ozcelik, A., Zheng, Y.B., and Huang, T.J., Acousto-Plasmofluidics: Acoustic Modulation of Surface Plasmon Resonance in Microfluidic Systems, AIP Advances, Vol. 5, pp. 097161, 2015.[PDF]

13. Wang, S.M., Zhao, C.L., Miao, X.Y., Zhao, Y.H., Rufo, J., Liu, Y.J., Huang, T.J., and Zheng, Y.B., Plasmofluidics: Merging Light and Fluids at the Micro-/Nano-Scale, Small, Vol. 11 (35), pp. 4423–4444, 2015.[PDF]

14. Yang, S.K., Slotcavage, D., Mai, J.D., Liang, W.S., Xie, Y.L., Chen, Y.C., and Huang, T.J., Combining the Masking and Scaffolding Modalities of Colloidal Crystal Templates: Plasmonic Nanoparticle Arrays with Multiple Periodicities, Chemistry of Materials, Vol. 26, pp. 6432−6438, 2014.[PDF]

15. Zhao, C.L., Liu, Y.M., Zhao, Y.H., Fang, N., and Huang, T.J., A Reconfigurable Plasmofluidic Lens, Nature Communications, Vol. 4, pp. 2305, 2013.[PDF]

16. Si*, G.Y., Zhao*, Y.H., Lv, J.T., Lu, M.Q., Wang, F.W.,  Liu, H.L., Xiang, N., Huang, T.J., Danner, A.J., Teng, J.H., and Liu, Y.J.,Reflective Plasmonic Color Filters Based on Lithographically Patterned Silver Nanorod Arrays, Nanoscale, Vol. 5, pp. 6243-6248, 2013.(*equal contributions)(featured as front cover image)[PDF]

17. Zhao, Y.H., Walker, T., Zheng, Y.B., Lin, S.S., Nawaz, A.A., Kiraly, B., Scott, J., and Huang, T.J., Mechanically Tuning the Localized Surface Plasmon Resonances of Gold Nanostructure Arrays, ASME Journal of Nanotechnology in Engineering and Medicine, Vol. 3, 011007, 2012.[PDF]

18. Zheng, Y.B., Kiraly, B., Weiss, P.S., and Huang, T.J., Molecular Plasmonics for Biology and Nanomedicine, Nanomedicine, Vol. 7(5),pp. 751-770, 2012.[PDF]

19. Zhao, Y.H., Hao, Q.Z., Ma, Y., Lu, M.Q., Zhang, B.X., Lapsley, M., Khoo, I.C., and Huang, T.J., Light-Driven Tunable Dual-Band Plasmonic Absorber using Liquid-Crystal-Coated Asymmetric Nanodisk Array, Applied Physics Letters, Vol. 100, pp. 053119, 2012. [PDF]

20. Lapsley, M.I., Shahravan,A., Hao, Q.Z., Juluri, B.K., Giardinelli, K., Lu, M.Q., Zhao, Y.H., Chiang, I.K., Matsoukas, T., and Huang, T.J., Shifts in Plasmon Resonance Due to Charging of A Nanodisk Array in Argon Plasma, Applied Physics Letters, Vol. 100, pp. 101903, 2012. (featured as front cover image) [PDF]

21. Juluri, B.K., Chaturvedi, N., Hao, Q.Z., Lu, M.Q., Velego, D., Jensen, L., and Huang, T.J., Scalable Manufacturing of Plasmonic Nanodisk Dimers and Cusp Nanostructures using Salting-out Quenching Method and Colloidal Lithography, ACS Nano, Vol. 5, pp. 5838–5847, 2011. [PDF]

22. Smalley, J.S.T., Zhao, Y.H., Nawaz, A.A., Hao, Q.Z., Ma, Y., Khoo, I.C., and Huang, T.J., High Contrast Modulation of Plasmonic Signals Using Nanoscale Dual-Frequency Liquid Crystals, Optics Express, Vol. 19, pp. 15265-15274, 2011. [PDF]

23. Zheng, Y.B., Kiraly, B., Cheunkar, S., Huang, T.J., and Weiss, P., Incident-Angle-Modulated Molecular Plasmonic Switches: A Case of Weak Exciton-Plasmon Coupling, Nano Letters, Vol. 11, pp. 2051-2055, 2011.[PDF]

24. Liu, Y.J., Zheng, Y.B., Liou, J., Chiang, I.K., Khoo, I.C., and Huang, T.J., All-Optical Modulation of Localized Surface Plasmon Coupling in a Hybrid System Composed of Photo-Switchable Gratings and Au Nanodisk Arrays, Journal of Physical Chemistry C, Vol. 115, pp. 7717–7722 2011. (featured as front cover image) [PDF]

25. Zhao, Y.H.,  Lin, S.S., Nawaz, A.A., Kiraly, B., Hao, Q.Z., Liu, Y.J. and Huang, T.J., Beam bending via plasmonic lenses, Optics Express, Vol. 18, pp. 23458-23465, 2010. [PDF]

26. Zheng*, Y.B., Juluri*, B.K., Jensen, L.L., Ahmed, D., Lu, M.Q., Jensen, L., and Huang, T.J., Dynamically Tuning Plasmon-Exciton Coupling in Arrays of Nanodisk-J-aggregate Complexes, Advanced Materials, Vol. 22, pp. 3603-3607, 2010. (*equal contributions) (featured as front cover image) [PDF]

27. Liu*, Y.J., Hao*, Q.Z., Smalley, J.S.T., Liou, J., Khoo, I.C., and Huang, T.J., A Frequency-Addressed Plasmonic Switch Based on Dual-Frequency Liquid Crystal, Applied Physics Letters, Vol. 97, pp. 091101, 2010. (*equal contributions) (featured as front cover image) [PDF]

28. Juluri, B.K., Lu, M.Q., Zheng, Y.B., Jensen, L., Huang, T. J., Coupling between Molecular and Plasmonic Resonances: Effect of Molecular Absorbance, Journal of Physical Chemistry C, Vol. 113, pp 18499–18503, 2009. [PDF]

29. Zheng, Y.B., Jensen, L.L., Yan, W., Walker, T.R., Juluri, B.K., Jensen, L., Huang, T. J., Chemically Tuning the Localized Surface Plasmon Resonances of Gold Nanostructure Arrays, Journal of Physical Chemistry C, Vol. 113, pp. 7019-7024, 2009. [PDF]

30. Mao, X.L., Lin, S.C.S., Lapsley, M.L., Shi, J.J., Juluri, B.K., and Huang, T.J., Tunable Liquid Gradient Refractive Index (L-GRIN) Lens with Two Degrees of Freedom,Lab on a Chip, Vol. 9, pp. 2050-2058, 2009. [PDF]

31. Juluri, B.K., Lin, S.S., Walker, T.R., Jensen, L. and Huang, T. J., Propagation of Designer Surface Plasmons in Structured Conductor Surfaces with Parabolic Gradient Index, Optics Express, Vol. 17, pp. 2997-3006, 2009. [PDF]

32.  Zheng, Y.B., Ying-Yan, W.g, Jensen, L., Fang, L., Juluri, B.K., Flood, A.H., Paul S Weiss, Stoddart, J.F., Huang, T. J., Active Molecular Plasmonics: Controlling Plasmon Resonances with Molecular Switches, Nano Letters, Vol. 9, pp. 819–825, 2009. [PDF]

33. Hsiao, V.K.S., Zheng, Y.B., Juluri, B.K., Huang, T.J., Light-Driven Plasmonic Switches Based on Au Nanodisk Arrays and Photoresponsive Liquid Crystals, Advanced Materials, Vol. 20, pp. 3528-3522, 2008. (featured as front cover image) [PDF]

34. Zheng, Y.B., Huang, T.J., Surface Plasmons of Metal Nanostructure Arrays: from Nanoengineering to Active Plasmonics, Journal of the Association for Laboratory Automation, Vol. 13, pp. 215-226, 2008. [PDF]

35. Juluri, B.K., Zheng, Y.B., Ahmed, D., Jensen, L., Huang, T.J., Effects of Geometry and Composition on Charge-Induced Plasmonic Shifts in Gold Nanoparticles, Journal of Physical Chemistry C, Vol. 112, pp. 7309-7312, 2008. [PDF]

36. Zheng, Y.B., Juluri, B.K., Mao, X.L., Walker, T.R., Huang, T.J., Systematic Investigation of Localized Surface Plasmon Resonance of Long-Range Ordered Au Nanodisk Arrays, Journal of Applied Physics, Vol. 103, pp. 014308, 2008. [PDF]

37.   Zheng, Y.B., Huang, T.J., Desai, A.Y., Wang, S.J., Tan, L.K., Gao, H., Huan, A.C.H., Thermal Behavior of Localized Surface Plasmon Resonance of Au/TiO2 Core/Shell Nanoparticle Arrays, Applied Physics Letters, Vol. 90, pp. 183117, 2007. [PDF]

38. Mao, X.L., Waldeisen, J.R., Juluri, B.K., Huang, T.J., Hydrodynamically Tunable Optofluidic Cylindrical Microlens, Lab on a Chip, Vol. 7, pp. 1303-1308, 2007. [PDF] 

Introduction

Since 2005, we have been working in the discipline of acoustofluidics (i.e., the fusion of micro- to nano-scale acoustics and fluid mechanics). Acoustofluidics has a combination of advantages that competing techniques lack: simple fabrication, high biocompatibility, versatility, compact and inexpensive devices and accessories, fast and effective fluid actuation, contact-free and non-invasive particle/cell manipulation, and compatibility with other lab-on-a-chip components. In addition, this discipline opens new avenues of research through the rare combinations of solids and fluids, mechanics and electronics, experiment and analysis, physics and materials, and engineering and biomedicine. Despite its promise, much remains to be explored in the field. Our objective in the next 5-10 years (Fig. 1) is to take the discipline to the next level and help translate acoustofluidic technologies from research labs to everyday use in real-world applications. To achieve this, many challenging problems related to physics, engineering, chemistry, and biomedical applications of acoustofluidics need to be overcome.

Image

Specifically, our research can be categorized into three topics: acoustic tweezers, sharp-edge-based acoustofluidics, bubble-based acoustofluidics, acoustofluidic-based MicroTAS, and optofluidics and plasmofluidics.

You can learn more about our acoustofluidics research through our Youtube channel: Acoustofluidics Lab.

Research Fields and Directions

We explore cutting-edge acoustofluidic technologies, advancing applications in biomedical engineering and precision medicine. Our current research focuses on manipulating cells and particles, developing multifunctional platforms for diagnostics, drug delivery, and tissue engineering. We utilize advanced acoustic and microfluidic systems to precisely control biological and fluidic interactions at the microscale, enabling innovative solutions for healthcare challenges.

Image			Acoustic Tweezers We develop innovative acoustofluidic techniques for precise manipulation of cells, particles, and fluids, through close collaborations with biologists and engineers at leading research institutions. Our interdisciplinary approach enables breakthroughs in biomedical applications such as diagnostics, therapeutic delivery, and tissue engineering.
Image			Acoustofluidics in biological sciences We leverage acoustofluidic technologies to address critical challenges in the biological sciences, focusing on cell sorting, tissue engineering, and molecular biology applications. Through collaborations with biologists and medical researchers, we aim to develop innovative tools for precision medicine, regenerative therapies, and cellular analysis, advancing healthcare and life science research.
Image			Acoustofluidic Detection and Disease Diagnosis We develop cutting-edge acoustofluidic technologies for the detection of biomarkers and rapid disease diagnosis. Collaborating with clinicians and biomedical engineers, we focus on creating highly sensitive, non-invasive diagnostic tools that can detect diseases at their earliest stages. Our work aims to revolutionize healthcare by enabling real-time, point-of-care diagnostics for improved patient outcomes.
Image			Droplet manipulation We develop advanced acoustofluidic platforms for precise droplet manipulation, enabling complex operations such as droplet generation, merging, and sorting. Collaborating closely with experts in biology and chemistry, we apply these technologies to enhance applications in diagnostics, drug delivery, and chemical analysis, providing versatile solutions for biomedical and chemical research.