High-speed Microfluidic Production of Phase-change Droplets for Gas Embolotherapy and as a Novel On-chip Pump

We report on a microfluidic device and droplet formation regime capable of generating clinical-scale quantities of liquid perfluoropentane droplets suitable in size and functionality for gas embolotherapy. We examine three modes of droplet formation in our flow-focusing device – geometry-controlled, dripping, and jetting – then use the dripping regime to generate highly monodisperse droplets in the range of 3-6 μm in diameter at rates exceeding 10 droplets per second. The phasechange droplets in this study are stable for weeks at room temperature yet undergo rapid liquid-to-gas phase transition, and volume expansion, above a uniform activation threshold. KEYWORDS: Droplet microfluidics, Droplet formation regimes, Clinical-scale, Phase-change, Occlusion, Cancer INTRODUCTION An accepted modality for the treatment of various cancers, embolization enables the reduction of blood flow into or out of a target vessel and has typically been used to produce local ischemia resulting in tumor necrosis [1]. In addition, the administration of chemotherapeutic agents alongside the embolic material has been shown to potentiate the treatment. Recently, the field of medical ultrasound has shown much interest in a novel form of embolization termed gas embolotherapy – in which acoustic or thermal energy induces a liquid perfluorocarbon droplet to undergo a liquid-to-gas phase transition and volume expansion to strategically form gas emboli in vivo. The phase change forms gas bubbles five to six times larger in diameter than the original droplets, enabling the intentional occlusion of such vessels as capillaries and some arterioles [2]. Current methods to generate these droplets, including sonication and high-speed mechanical agitation, typically result in a polydisperse size distribution and a non-uniform acoustic activation threshold, as well as in droplets either too large for passage through the lungs or too small to occlude the desired vessel [2,3]. Though droplet-based microfluidic systems have shown to be effective technologies to produce liquid-in-liquid droplet emulsions in the micrometer diameter range with high uniformity, these systems have been viewed as unviable as bulk manufacturing processes owing to low generation rates and inappropriate emulsion characteristics. We report on a microfluidic flow-focusing device operated in the dripping regime capable of generating liquid perfluoropentane droplets in the range of 3-6 μm in diameter at rates exceeding 10 droplets per second. Resulting droplet populations exhibit polydispersity index values of <5%, are stable at room temperature, and are thermally vaporized at 88C in a water bath. Further, we introduce a novel use for these phase-change droplet populations as an on-chip pump, activated by droplet heating, to deliver reagents from a storage reservoir to a new location on the chip. THEORY Recent publications have reported the observation of a number of distinct droplet formation regimes, including geometry-controlled, dripping, and jetting [4,5]. While droplets in the geometry-controlled mode are limited in size by the width of the smallest feature in the microfluidic device, the orifice, droplets in the dripping mode of droplet breakup may be smaller and are generated with a shorter primary breakup period than other modes [4]. We thus set out to use the dripping mode to generate mass quantities of liquid perfluoropentane droplets in the appropriate size range for gas embolotherapy as a cancer treatment. EXPERIMENTAL The microfluidic device, made of PDMS on a glass substrate and pictured in Figure 1, consists of fixed geometric channels designed to direct liquid perfluoropentane and lipid solution (an aqueous glycerol mixture with the stabilizing lipids DSPC and DSPE-PEG2000 in a 9:1 molar ratio) to an orifice 7 μm in width. Standard soft lithography techniques were used to fabricate the microfluidic devices. A lipid shell was chosen over an albumin shell, as albumin shells have shown an increased tendency to lodge in the pulmonary capillaries of the patient [3]. Perfluoropentane (dodecafluoropentane, C5F12) was chosen as the liquid perfluorocarbon. Pressure pumping using an in-house setup was used to increase stability in the liquid perfluoropentane flow. 978-0-9798064-4-5/μTAS 2011/$20©11CBMS-0001 79 15th International Conference on Miniaturized Systems for Chemistry and Life Sciences October 2-6, 2011, Seattle, Washington, USA Droplets were collected in the outlet well of the microfluidic device and stored in a sealed glass vial at room temperature to simulate an on-the-shelf setting. ImageJ was used to assess the stability of droplets over a two-week period. To determine the vaporization temperature of the phase-change droplets, aliquots of droplet solution were added to dilute lipid solution in a vented glass vial. The vial was suspended in a stirred water bath and the bath temperature was slowly raised until observation of gas bubble formation. RESULTS AND DISCUSSION By adjusting the flow parameters in our flow-focusing device we observed three distinct droplet formation regimes. A protrudeand-retract mechanism of droplet formation characterizes the geometry-controlled mode and limits the droplet diameter to the width of the orifice. Droplets generated in the geometry-controlled mode ranged from 7.9 ± .1 μm at 2.44x10 Hz to 10.8 ± .2 μm at 3.28x10 Hz. Increasing the continuous phase flow, and consequently the capillary number, transitions the device into the dripping regime. The dispersed phase finger highly narrows and does not retract, but rather remains at a fixed location in the orifice, and droplets break off due to Rayleigh capillary instability. Diameters of droplets generated in the dripping regime ranged from 3.6 ± .2 μm at 1.36x10 Hz to 10.8 ± .1 μm at 5.00x10 Hz. Further increasing the capillary number transitions the device into the jetting regime, characterized by a dispersed phase finger which extends far beyond the orifice and again experiences Rayleigh capillary instability. The flow rate parameters play a determining role in the stability of production as well as the size and generation frequency of the liquid perfluoropentane droplets. In general, droplet diameter D increases with the dispersed phase pressure PP for a fixed continuous phase flow QL. Whereas the droplet generation frequency fd increases with PP for a fixed QL in geometry-controlled mode to conserve mass, the opposite is true in the dripping regime: for a given QL in the dripping regime, decreasing PP significantly quickens the generation frequency while reducing D. Shown in Figure 2 is the effect of the unconverted flow rate ratio – defined as α = PP/QL where PP represents the liquid perfluoropentane pressure and QL represents the flow of the lipid solution – on the droplet generation rate and size in the dripping regime. A decrease in α tends to increase the generation rate of droplets and decrease their size. Thus, increasing the continuous outer phase flow relative to the dispersed inner phase flow in the dripping regime enables the super high-speed generation of droplets suitable in size for therapeutic applications. Figure 3 shows an image from high-speed video of droplet production in the dripping regime. In this regime the dispersed phase finger highly narrows and droplets break off due to Rayleigh capillary instability at sizes approximating the width of the dispersed phase tip. We assessed a population of droplets, with a mean diameter of 4.5 μm, for stability over time and observed a drift in diameter of less than 4% over two weeks at room temperature. Pure liquid perfluoropentane has a boiling point of 29C, suggesting spontaneous liquid-to-gas transition at body temperature. However, as with previous studies, encapsulation of the liquid in a lipid shell increases Figure 1: (a) Schematic of the microfluidic flowfocusing device. (b) Photograph of the PDMS device next to a dime, for size comparison. (c) Main functional area of the device with a 7 μm orifice. Direction of flow is indicated by the arrows. liquid perfluoropentane