Cell separation using tilted-angle standing surface acoustic waves

Separation of cells is a critical process for studying cell properties, disease diagnostics, and therapeutics. Cell sorting by acoustic waves offers a means to separate cells on the basis of their size and physical properties in a label-free, contactless, and biocompatible manner. The separation sensitivity and efficiency of currently available acoustic-based approaches, however, are limited, thereby restricting their widespread application in research and health diagnostics. In this work, we introduce a unique configuration of tilted-angle standing surface acoustic waves (taSSAW), which are oriented at an optimally designed inclination to the flow direction in the microfluidic channel. We demonstrate that this design significantly improves the efficiency and sensitivity of acoustic separation techniques. To optimize our device design, we carried out systematic simulations of cell trajectories, matching closely with experimental results. Using numerically optimized design of taSSAW, we successfully separated 2- and 10-µm-diameter polystyrene beads with a separation efficiency of ~99%, and separated 7.3- and 9.9-µm-polystyrene beads with an efficiency of ~97%. We illustrate that taSSAW is capable of effectively separating particles–cells of approximately the same size and density but different compressibility. Finally, we demonstrate the effectiveness of the present technique for biological–biomedical applications by sorting MCF-7 human breast cancer cells from nonmalignant leukocytes, while preserving the integrity of the separated cells. The method introduced here thus offers a unique route for separating circulating tumor cells, and for label-free cell separation with potential applications in biological research, disease diagnostics, and clinical practice.National Institutes of Health (U.S.) (Grant U01HL114476)National Institutes of Health (U.S.) (New Innovator Award 1DP2OD007209-01)National Science Foundation (U.S.). Materials Research Science and Engineering Centers (Program) (Grant DMR-0820404

Rapid cell separation with minimal manipulation for autologous cell therapies

The ability to isolate specific, viable cell populations from mixed ensembles with minimal manipulation and within intra-operative time would provide significant advantages for autologous, cell-based therapies in regenerative medicine. Current cell-enrichment technologies are either slow, lack specificity and/or require labelling. Thus a rapid, label-free separation technology that does not affect cell functionality, viability or phenotype is highly desirable. Here, we demonstrate separation of viable from non-viable human stromal cells using remote dielectrophoresis, in which an electric field is coupled into a microfluidic channel using shear-horizontal surface acoustic waves, producing an array of virtual electrodes within the channel. This allows high-throughput dielectrophoretic cell separation in high conductivity, physiological-like fluids, overcoming the limitations of conventional dielectrophoresis. We demonstrate viable/non-viable separation efficacy of > 98% in pre-purified mesenchymal stromal cells, extracted from human dental pulp, with no adverse effects on cell viability, or on their subsequent osteogenic capabilities

Acoustic separation of circulating tumor cells

Circulating tumor cells (CTCs) are important targets for cancer biology studies. To further elucidate the role of CTCs in cancer metastasis and prognosis, effective methods for isolating extremely rare tumor cells from peripheral blood must be developed. Acoustic-based methods, which are known to preserve the integrity, functionality, and viability of biological cells using label-free and contact-free sorting, have thus far not been successfully developed to isolate rare CTCs using clinical samples from cancer patients owing to technical constraints, insufficient throughput, and lack of long-term device stability. In this work, we demonstrate the development of an acoustic-based microfluidic device that is capable of high-throughput separation of CTCs from peripheral blood samples obtained from cancer patients. Our method uses tilted-angle standing surface acoustic waves. Parametric numerical simulations were performed to design optimum device geometry, tilt angle, and cell throughput that is more than 20 times higher than previously possible for such devices. We first validated the capability of this device by successfully separating low concentrations (~100 cells/mL) of a variety of cancer cells from cell culture lines from WBCs with a recovery rate better than 83%. We then demonstrated the isolation of CTCs in blood samples obtained from patients with breast cancer. Our acoustic-based separation method thus offers the potential to serve as an invaluable supplemental tool in cancer research, diagnostics, drug efficacy assessment, and therapeutics owing to its excellent biocompatibility, simple design, and label-free automated operation while offering the capability to isolate rare CTCs in a viable state.National Institutes of Health (U.S.) (Grant 1 R01 GM112048-01A1)National Institutes of Health (U.S.) (Grant 1R33EB019785-01)National Science Foundation (U.S.)Penn State Center for Nanoscale Science (Materials Research Science and Engineering Center Grant DMR-0820404)National Institutes of Health (U.S.) (Grant U01HL114476

Rare-cell enrichment by a rapid, label-free, ultrasonic isopycnic technique for medical diagnostics

One significant challenge in medical diagnostics lies in the development of label-free methods to separate different cells within complex biological samples. Here we demonstrate a generic, low-power ultrasonic separation technique, able to enrich different cell types based upon their physical properties. For malaria, we differentiate between infected and non-infected red blood cells in a fingerprick-sized drop of blood. We are able to achieve an enrichment of circulating cells infected by the ring stage of the parasite over nonparasitized red blood cells by between two and three orders of magnitude in less than 3 seconds (enabling detection at parasitemia levels as low as 0.0005 %). In a second example, we also show that our methods can be used to enrich different cell types, concentrating Trypanosoma in blood at very low levels of infection, on disposable, low-cost chips

Particle separation by phase modulated surface acoustic waves

High efficiency isolation of cells or particles from a heterogeneous mixture is a critical processing step in lab-on-a-chip devices. Acoustic techniques offer contactless and label-free manipulation, preserve viability of biological cells, and provide versatility as the applied electrical signal can be adapted to various scenarios. Conventional acoustic separation methods use time-of-flight and achieve separation up to distances of quarter wavelength with limited separation power due to slow gradients in the force. The method proposed here allows separation by half of the wavelength and can be extended by repeating the modulation pattern and can ensure maximum force acting on the particles. In this work, we propose an optimised phase modulation scheme for particle separation in a surface acoustic wave microfluidic device. An expression for the acoustic radiation force arising from the interaction between acoustic waves in the fluid was derived. We demonstrated, for the first time, that the expression of the acoustic radiation force differs in surface acoustic wave and bulk devices, due to the presence of a geometric scaling factor. Two phase modulation schemes are investigated theoretically and experimentally. Theoretical findings were experimentally validated for different mixtures of polystyrene particles confirming that the method offers high selectivity. A Monte-Carlo simulation enabled us to assess performance in real situations, including the effects of particle size variation and non-uniform acoustic field on sorting efficiency and purity, validating the ability to separate particles with high purity and high resolution

Cell detachment and label-free cell sorting using modulated surface acoustic waves (SAW) in droplet-based microfluidics

We present a droplet-based surface acoustic wave (SAW) system designed to viably detach biological cells from a surface and sort cell types based on differences in adhesion strength (adhesion contrast), without the need to label cells with molecular markers. The system uses modulated SAW to generate pulsatile flows in the droplets and efficiently detach the cells, thereby minimizing SAW excitation power and exposure time. As a proof-of-principle, the system is shown to efficiently sort HEK 293 from A7r5 cells based on adhesion contrast. Results are obtained in minutes with sorting purity and efficiency reaching 97 % and 95 %, respectively.Comment: Accepted for publication in Lab on a Chi

Direct Observation of Propagating Gigahertz Coherent Guided Acoustic Phonons in Free Standing Single Copper Nanowires

We report on gigahertz acoustic phonon waveguiding in free-standing single copper nanowires studied by femtosecond transient reflectivity measurements. The results are discussed on the basis of the semianalytical resolution of the Pochhammer and Chree equation. The spreading of the generated Gaussian wave packet of two different modes is derived analytically and compared with the observed oscillations of the sample reflectivity. These experiments provide a unique way to independently obtain geometrical and material characterization. This direct observation of coherent guided acoustic phonons in a single nano-object is also the first step toward nanolateral size acoustic transducer and comprehensive studies of the thermal properties of nanowires

Whole-blood sorting, enrichment and in situ immunolabeling of cellular subsets using acoustic microstreaming

Analyzing undiluted whole human blood is a challenge due to its complex composition of hematopoietic cellular populations, nucleic acids, metabolites, and proteins. We present a novel multi-functional microfluidic acoustic streaming platform that enables sorting, enrichment and in situ identification of cellular subsets from whole blood. This single device platform, based on lateral cavity acoustic transducers (LCAT), enables (1) the sorting of undiluted donor whole blood into its cellular subsets (platelets, RBCs, and WBCs), (2) the enrichment and retrieval of breast cancer cells (MCF-7) spiked in donor whole blood at rare cell relevant concentrations (10 mL− 1), and (3) on-chip immunofluorescent labeling for the detection of specific target cellular populations by their known marker expression patterns. Our approach thus demonstrates a compact system that integrates upstream sample processing with downstream separation/enrichment, to carry out multi-parametric cell analysis for blood-based diagnosis and liquid biopsy blood sampling

Three-Dimensional Numerical Modeling of Acoustic Trapping in Glass Capillaries

Acoustic traps are used to capture and handle suspended microparticles and cells in microfluidic applications. A particular simple and much-used acoustic trap consists of a commercially available, millimeter-sized, liquid-filled glass capillary actuated by a piezoelectric transducer. Here, we present a three-dimensional numerical model of the acoustic pressure field in the liquid coupled to the displacement field of the glass wall, taking into account mixed standing and traveling waves as well as absorption. The model predicts resonance modes well suited for acoustic trapping, their frequencies and quality factors, the magnitude of the acoustic radiation force on a single test particle as a function of position, and the resulting acoustic retention force of the trap. We show that the model predictions are in agreement with published experimental results, and we discuss how improved and more stable acoustic trapping modes might be obtained using the model as a design tool.Comment: 13 pages, 15 pdf figures, pdfLatex/Revte