Scaleable production of microbubbles using an ultrasound-modulated microfluidic device.

Surfactant-coated gas microbubbles are widely used as contrast agents in ultrasound imaging and increasingly in therapeutic applications. The response of microbubbles to ultrasound can be strongly influenced by their size and coating properties, and hence the production method. Ultrasonic emulsification (sonication) is the most commonly employed method and can generate high concentrations of microbubbles rapidly, but with a broad size distribution, and there is a risk of contamination and/or degradation of sensitive components. Microfluidic devices provide excellent control over microbubble size, but are often challenging or costly to manufacture, offer low production rates (<106s-1), and are prone to clogging. In this study, a hybrid sonication-microfluidic or "sonofluidic" device was developed. Bubbles of ∼180 µm diameter were produced rapidly in a T-junction and subsequently exposed to ultrasound (71-73 kHz) within a microchannel, generating microbubbles (mean diameter: 1-2 µm) at a rate of >108s-1 using a single device. Microbubbles were prepared using either the sonofluidic device or conventional sonication, and their size, concentration, and stability were comparable. The mean diameter, concentration, and stability were found to be comparable between techniques, but the microbubbles produced by the sonofluidic device were all <5 µm in diameter and thus did not require any post-production fractionation.

Generation of functional hepatocyte 3D discoids in an acoustofluidic bioreactor.

Ultrasonic standing wave systems have previously been used for the generation of 3D constructs for a range of cell types. In the present study, we cultured cells from the human hepatoma Huh7 cell line in a Bulk Acoustic Wave field and studied their viability, their functions, and their response to the anti-cancer drug, 5 Fluorouracil (5FU). We found that cells grown in the acoustofluidic bioreactor (AFB) expressed no reduction in viability up to 6 h of exposure compared to those cultured in a conventional 2D system. In addition, constructs created in the AFB and subsequently cultured outside of it had improved functionality including higher albumin and urea production than 2D or pellet cultures. The viability of Huh7 cells grown in the ultrasound field to 5FU anti-cancer drug was comparable to that of cells cultured in the 2D system, showing rapid diffusion into the aggregate core. We have shown that AFB formed 3D cell constructs have improved functionality over the conventional 2D monolayer and could be a promising model for anti-cancer drug testing.

Acoustically modulated biomechanical stimulation for human cartilage tissue engineering.

Bioacoustofluidics can be used to trap and levitate cells within a fluid channel, thereby facilitating scaffold-free tissue engineering in a 3D environment. In the present study, we have designed and characterised an acoustofluidic bioreactor platform, which applies acoustic forces to mechanically stimulate aggregates of human articular chondrocytes in long-term levitated culture. By varying the acoustic parameters (amplitude, frequency sweep, and sweep repetition rate), cells were stimulated by oscillatory fluid shear stresses, which were dynamically modulated at different sweep repetition rates (1-50 Hz). Furthermore, in combination with appropriate biochemical cues, the acoustic stimulation was tuned to engineer human cartilage constructs with structural and mechanical properties comparable to those of native human cartilage, as assessed by immunohistology and nano-indentation, respectively. The findings of this study demonstrate the capability of acoustofluidics to provide a tuneable biomechanical force for the culture and development of hyaline-like human cartilage constructs in vitro.

A thin-reflector microfluidic resonator for continuous-flow concentration of microorganisms: a new approach to water quality analysis using acoustofluidics.

An acoustofluidic device has been developed for concentrating vegetative bacteria in a continuous-flow format. We show that it is possible to overcome the disruptive effects of acoustic streaming which typically dominate for small target particles, and demonstrate flow rates compatible with the testing of drinking water. The device consists of a thin-reflector multi-layered resonator, in which bacteria in suspension are levitated towards a glass surface under the action of acoustic radiation forces. In order to achieve robust device performance over long-term operation, functional tests have been carried out to (i) maintain device integrity over time and stabilise its resonance frequency, (ii) optimise the operational acoustic parameters, and (iii) minimise bacterial adhesion on the inner surfaces. Using the developed device, a significant increase in bacterial concentration has been achieved, up to a maximum of ~60-fold. The concentration performance of thin-reflector resonators was found to be superior to comparable half-wave resonators.