Controllable in-situ cell electroporation with cell positioning and impedance monitoring using micro electrode array

Abstract

This paper reports a novel microarray chip for in-situ, real-time and selective electroporation on individual cells integrated with cell positioning and impedance monitoring. An array of quadrupole-electrode units (termed positioning electrodes) and pairs of planar center electrodes located at the centers of each quadrupole-electrode unit were fabricated on the chip. The positioning electrodes are used to trap and position living cells onto the center electrodes based on negative dielectrophoresis (nDEP). The center electrodes are used for in-situ cell electroporation, and also used to measure cell impedance for monitoring cellular dynamics in real time. Controllably selective electroporation and electrical measurement on the cells in array are realized. We present an evidence of selective electroporation through use of fluorescent dyes. Subsequently we use in-situ and real-time impedance measurement to monitor the process, which demonstrates the dynamic behavior of the cell electroporation. Finally, we show the use of this device to perform successful transfection onto individual HeLa cells with vector DNA encoding a green fluorescent.

Figure 1. Structure of the device.

(a) Electrode array. (b) Structure of one quadrupole-electrode unit with a pair of center electrodes located at the center and the inset shows the surface morphology of the center electrodes. (c) The chip is assembled onto the PCB. (d) A PDMS pool with the inlet and the outlet of the microfluidic is adhered onto the chip to form a cell sample pool. (e) The prototype of the device.

Figure 2. Experimental setup.

It consists of a fluorescent microscope, a syringe pump that pumps sample into the pool of the device, and a main board integrating controlling circuit and I/O ports which are connected to a function generator and an impedance analyzer. The syringe pump, function generator and impedance analyzer are controlled by using a computer.

Figure 3. The process of cell positioning, electroporation, and impedance measurement.

(a) Cell is positioned at the unit center under nDEP. (b) The electroporation is conducted onto the positioned cell using two center electrodes. (c) The impedance measurement is conducted to monitor the cellular dynamic.

Figure 4. Selective control for cell electroporation.

(a) Schematic diagram of electrical I/O connection of the center electrodes for electroporating the cells in a line of the array. (b) The cells are positioned on the measuring electrodes to form an array by nDEP. (c,d) Demonstrations of selective electroporation in array dyed by PI.

Figure 5. Electroporation analysis using fluorescent dyes.

(a) HeLa cells subjected to different electroporation voltage correspond different PI and Calcein-AM fluorescence intensity. Error bars indicate statistic characteristics of fluorescence intensity in one chip. (b) The fluorescence intensity of PI and Calcein-AM corresponding to different pulse width of electroporation signal. (c) The distribution of the electric field intensity within one unit under an electroporation voltage of 10 V.

Figure 6. The single cellular impedances at 100 kHz before and after electroporation.

Figure 7. The cell transfection result.

The red circles indicate sites where the center electrodes locate. The successfully transfected cells fluoresce green.