Acoustic Trapping and Manipulation of Hollow Microparticles under Fluid Flow Using a Single-Lens Focused Ultrasound Transducer

Abstract

Microparticle manipulation and trapping play pivotal roles in biotechnology. To achieve effective manipulation within fluidic flow conditions and confined spaces, it is necessary to consider the physical properties of microparticles and the types of trapping forces applied. While acoustic waves have shown potential for manipulating microparticles, the existing setups involve complex actuation mechanisms and unstable microbubbles. Consequently, the need persists for an easily deployable acoustic actuation setup with stable microparticles. Here, we propose the use of hollow borosilicate microparticles possessing a rigid thin shell, which can be efficiently trapped and manipulated using a single-lens focused ultrasound (FUS) transducer under physiologically relevant flow conditions. These hollow microparticles offer stability and advantageous acoustic properties. They can be scaled up and mass-produced, making them suitable for systemic delivery. Our research demonstrates the successful trapping dynamics of FUS within circular tubings of varying diameters, validating the effectiveness of the method under realistic flow rates and ultrasound amplitudes. We also showcase the ability to remove hollow microparticles by steering the FUS transducer against the flow. Furthermore, we present potential biomedical applications, such as active cell tagging and navigation in bifurcated channels as well as ultrasound imaging in mouse cadaver liver tissue.

Keywords: acoustic manipulation; acoustic trapping; focused ultrasound; hollow microparticles; microbubbles; microrobotics; particle manipulation; ultrasound imaging.

Figure 1

Schematic description of a single-lens FUS transducer trapping hollow particles. The particles (blue) are trapped inside a bifurcated tubing against fluid flow. The single-lens FUS waves used to trap the particles is placed below the tubing. The transmitted focused traveling ultrasound waves apply acoustic trapping force to create a stable aggregate near the focal spot.

Figure 2

Characterization of the hollow particles and acoustic setup. a) Schematic representation of a hollow core–shell particle with inner radius r_i and outer radius r_o. b) Light microscope image (left) and scanning electron microscope (SEM) image (right) of polydisperse hollow particles. c) This graph shows the acoustophoretic contrast factor for different ratios α of r_i and r_o. For small α, as in case of a solid particles, ϕ is positive. When increasing the ratio α the acoustophoretic contrast factor gets highly negative inverting the acoustophoretic behavior of the hollow particles as used for this study compared to solid particles. d) Representation of the acoustic radiation force dependency on the radius of a particle and the ratio α. The graph shows that the radiation force acting on solid particles is positive while it is negative for hollow particles. All values for graphs g) and h) are obtained by numerical simulations. e) COMSOL finite element simulation of the pressure distribution around a hollow core–shell particle immersed into a standing acoustic wave. f) Finite element simulated pressure applied by a 2 MHz FUS transducer into a 500 μm diameter tubing made from Tygon.

Figure 3

Trapping dynamics of the hollow particles inside a FUS field. a) Schematic depiction of the experimental setup used to evaluate the trapping efficacy of acoustic traveling wave-induced agglomeration of hollow particles inside a 500 μm diameter Tygon tubing. A FUS transducer with a central frequency of 2 MHz is placed inside a water tank. A tubing with a syringe pump was positioned under water using a manual x-y-z stage. The hollow particles were flushed into the tubing using the syringe pump. The liquid was stored in a reservoir. b) Microscope image showing the particles flowing through a Tygon tubing before (top) and after (bottom) applying the FUS. Using a customized Python code different region are defined for automated particle concentrations measurements. The blue dashed line named L1 indicates the cross section of the tubing before the FUS focal spot. The yellow dashed line A_trap and the blue dashed line L2 indicate the trapping region and after trapping region in the flow direction, respectively. c) Time history images of the pixel intensity of the L1 (top) and L2 (bottom) cross section before and after applying the FUS. Both b) and c) refer to an applied flow speed of 25.4 mm/s and a FUS input voltage of 200 mV (peak-to-peak). d) This plot shows the normalized particle concentration inside the trapping region A_trap over time for flow rates reaching from 0.1 to 0.7 mL/min corresponding to the flow speeds of 8.5–59.4 mm/s.

Figure 4

Control and retrieval of the hollow particles against the fluid flow. a) Schematic representation of hollow particles retrieved using acoustic manipulation enabled by a FUS transducer against the fluid flow. b) Time lapse images showing the retrieval of the hollow microparticle swarm immersed into a 4.77 mm-diameter Tygon tubing against a flow speed of 0.93 mm/s. The particles are trapped by using a 500 kHz focused ultrasound transducer. The particles are moved back to the injection side by using a relative movement between the tubing and the transducers. There they are retrieved using the same 28 G needle used for injection. c) Movement of acoustically trapped hollow microparticles inside a 500 μm diameter tubing. Here 100 μL of particles with a concentration of 5 mg/mL are injected into a tubing using a 28 G needle. After injection, the particles are trapped using a 2 MHz focused ultrasound transducer and moved inside the tubing. The particles are imaged using a high-speed camera mounted to a light microscope. The applied flow speed is 25.46 mm/s. d) The particles are trapped using a focused 500 kHz transducer. Using a manual x-y-z stage the particles were moved inside the tubing in 2D. The applied flow speed was 4.67 mm/s. For all figures, the trajectories of the particles are marked in blue, and the flow direction is marked in orange. e) Ultrasonographic images of needle positioning (left) and subsequent particle injection (right) into the portal vein of the liver of an ex vivo mouse with multiple liver metastasis. The liver area is marked in yellow. Ultrasound imaging is performed in the B-mode with a frequency of 40 MHz.

Figure 5

Trapped particle swarm movement inside a bifurcated microfluidic channel and active cell tagging. a) Schematic representation of the functionalization procedure for binding an anti-HER2 antibody to hollow borosilicate particles. First NH2 groups are added to the particles using APTES modification. Subsequently NHS-Biotin and streptavidin are bound to the particles. In the last step, a biotinylated anti-HER2 is bound to the particles using streptavidin–biotin interaction. b) Movement of hollow particles inside a bifurcated microfluidic chip intended for cell culture. Here a flow rate of 0.75 mL/min was applied. The flow direction is depicted by a green arrow. From the starting position t₀ the particles are moved in the top bifurcation against the flow. Afterward the particles were moved back to the starting position in the direction of the flow. Finally, the particles were moved to the bottom bifurcation and back to the starting point. For acoustic trapping, a focused 2 MHz transducer was used. The current trapping region is marked in each image by a blue dotted circle. c) Microscope images showing the trapping and subsequent movement of hollow particles through a microfluidic channel toward a tumor spheroid (yellow frame). The trapping region is indicated by a purple dotted circle. When near the tumor spheroid the particles attached to the tumor spheroid (pink frame) and escaped the acoustic trap (t = 5.93 s). The color table of the microscope images was changed to green for better visibility. d) Light microscope image of SKBR3 cells (immortalized human breast cancer cell line) embedded into a cell culture chip surrounded by a yellow frame. The image surrounded by a pink frame is taken after injection and FUS-enabled movement of anti-HER2 functionalized hollow particles to the cancer spheroid. The particles are bound to the cells by an antibody-receptor interaction.