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Electrosonic MEMS Gun for Efficient Cellular Transfection and Drug Delivery

Vladimir G. Zarnitsyn1, F. Levent Degertekin2, and Andrei G. Fedorov1. (1) George W. Woodruff School of Mechanical Enginnering, Georgia Institute of Technology, 801 Ferst Drive, Atlanta, GA 30332-0405, (2) George W. Woodruff School of Mechanical Engineering, Georgia Institute of Technology, 801 Ferst Drive, Atlanta, GA 30332-0405

We developed the electrosonic MEMS gun for on-demand drug/gene delivery into mammalian cells via combination of ultrasonic (mechanical) and electrical poration of the cell membrane. In addition to poration multi-functionality, the device has capability for in-line size selective cell sorting, and it also enables transport of transfected cells as a post-transfection ejection step for further processing. The critical advantage of this technology is the uniformity of the mechano/electro-poration action experienced by each cell individually, thereby enabling high degree of transfection/delivery control and efficiency. A device also minimizes the required sample size (~100 nL), improves sample utilization, and is inherently suitable for parallel, high throughput operation as well as multiplexing in the array format. Further, the electrosonic MEMS gun technology is inexpensive enough to be made disposable since the devices are batch microfabricated in silicon using a simple, well reproducible process. A device is designed to work in an array format, so it can operate in both the high throughput mode and also in the multiplexed mode if the array is divided into individually controlled compartments. Each compartment is loaded with a buffer solution, suspension of biological cells of one or several types and dimensions, and foreign biomolecules (e.g., DNA, RNA, proteins, etc) that one desires to inject into the cells. The micromachined liquid horn nozzle structures of the loaded chambers efficiently focus ultrasonic waves generated by the piezoelectric transducer at one of the resonant frequencies of the fluid cavities, leading to establishment of the local pressure gradient near the tip/opening of the nozzles. This pressure gradient at the nozzle tip serves two important functions: (1) it allows to eject on-demand droplets from the device, and (2) it allows to apply a mechanical (pressure) force of controlled magnitude and duration to the cell membrane as each cell passes through the nozzle neck during the ejection leading to membrane poration and injection of extracellular biomolecules (DNA, RNA, proteins) from solution into the cell through open pores. Efficient sonoporation occurs when amplitude of the acoustic pressure pulse applied to the cell membrane is between 1 and 100 kPa (in access of the DC hydrostatic pressure) and the pulse duration in the range of 0.1 and 10 μs. These operating parameters are readily realized in operation of the electrosonic DNA gun by varying an amplitude and frequency of electric signal applied to the piezoelectric transducer. Mechanical stress generated by ultrasound has been shown to induce temporal cell permeabilization to drugs and naked DNA with subsequent cell transfection. Our experimental results show low power (<100 mW) and temperature (<30 oC) transfection without device clogging by biomolecules/cells and with proven thermal stability of operation. Comparison of flow cytometry results unambiguously indicates that biological cells remain alive upon ejection by the electrosonic MEMS gun. Further, we experimentally demonstrate efficient uptake of calcein into cells upon processing by the device. Thus, use of the electrosonic gun enables cell treatment which has drug (possibly DNA/gene) delivery potential. Acknowledgement: we would like to thank Prof. Andres Garcia and J. Mark Meacham for use of the experimental facilities and help with experiments.