Therapeutic approaches that enhance thrombolysis by combining recombinant tissue plasminogen activator (rtPA), ultrasound, and/or microbubbles (MBs) are known as sonothrombolysis techniques. To date, sonothrombolysis approaches have primarily utilized commercially available MB formulations (or derivatives thereof) with diameters in the range 1-4 µm and circulation lifetimes between 5 and 15 min. The present study evaluated the in vitro sonothrombolysis efficacy of large diameter MBs (d ≥ 10 µm) with much shorter lifetimes that were produced on demand and in close proximity to the blood clot using a flow-focusing microfluidic device. MBs with a N gas core and a non-crosslinked bovine serum albumin shell were produced with diameters between 10 and 20 µm at rates between 50 and 950 × 10 per second. Use of these large MBs resulted in approximately 4.0-8.8 fold increases in thrombolysis rates compared to a clinical rtPA dose and approximately 2.1-4.2 fold increases in thrombolysis rates compared to sonothrombolysis techniques using conventional MBs. The results of this study indicate that the large diameter microbubbles with transient stability are capable of significantly enhanced in vitro sonothrombolysis rates when delivered directly to the clot immediately following production by a flow focusing microfluidic device placed essentially in situ adjacent to the clot.

Flow focusing microfluidic devices (FFMDs) have been investigated for the production of monodisperse populations of microbubbles for chemical, biomedical and mechanical engineering applications. High-speed optical microscopy is commonly used to monitor FFMD microbubble production parameters, such as diameter and production rate, but this limits the scalability and portability of the approach. In this work, a novel FFMD design featuring integrated electronics for measuring microbubble diameters and production rates is presented. A micro Coulter particle counter (μCPC), using electrodes integrated within the expanding nozzle of an FFMD (FFMD-μCPC), was designed, fabricated and tested. Finite element analysis (FEA) of optimal electrode geometry was performed and validated with experimental data. Electrical data was collected for 8-20 μm diameter microbubbles at production rates up to 3.25 × 105 MB s-1 and compared to both high-speed microscopy data and FEA simulations. Within a valid operating regime, Coulter counts of microbubble production rates matched optical reference values. The Coulter method agreed with the optical reference method in evaluating the microbubble diameter to a coefficient of determination of R2 = 0.91.

Real-time observation and control of particle size and production rate in microfluidic devices are important capabilities for a number of applications, including the production, sorting, and manipulation of microbubbles and droplets. The production of microbubbles from flow-focusing microfluidic devices had been investigated in multiple studies, but each lacked an approach for on-chip measurement and control of microbubble diameter in real time. In this work, we implement a closed-loop feedback control system in a flow-focusing microfluidic device with integrated on-chip electrodes. Using our system, we measure and count microbubbles between 13 and 28 μm in diameter and control their diameter using a proportional–integral controller. We validate our measurements against an optical benchmark with R2=0.98 and achieve a maximum production rate of 1.4×105/s. Using the feedback control system, the device enabled control in microbubble diameter over the range of 14–24 μm.