(Invited) Discrimination and Selection of Target Cells from the Cell-Based Array Based on Dielectrophoresis

Abstract

Introduction: We have developed manipulation techniques to form cell-based arrays by positive dielectrophoresis (p-DEP) and to retrieve target cells from cell−based arrays selectively by negative dielectrophoresis (n-DEP). The novel devices with microwell arrays on microband electrodes were employed to manipulate cells. Hybridomas with the secretion ability of antibodies were trapped to form cell-based arrays. Then, the ability was discriminated by trapping the antibodies to the antigen immobilized on the bottom of wells. Finally, individual cells trapped in microwells were selectively retrieved by regulating the generation of electric fields in individual microwells. The development of series of these techniques could be useful to recover hybridomas producing antibodies with high affinity in large populations of cells without repeated steps of a culture and a limiting serial-dilution. Experimental: The device comprises the upper substrate with indium-tin-oxide (ITO) microband electrodes and the lower substrate with microwell arrays on ITO microband electrodes. Hybridomas producing an anti-rabbit serum albumin (RSA) antibody suspended in the DEP medium were introduced in the channel. AC signal (3 MHz, 2 Vpp) was then applied to the upper and lower microband electrodes with opposite phase to form a cell-based array. Anti-RSA antibodies secreted from hybridomas trapped in the microwells were captured by RSA immobilized on the electrodes. Cell arrays were then treated with anti-mouse IgG antibody conjugated with Alexa 488 (a secondary antibody). Thereafter, a repulsive force of negative dielectrophoresis (n-DEP) was employed to retrieve the target hybridomas from the microwell array. To retrieve the target hybridoma, an AC signal in the n-DEP frequency region was applied to a pair of microband electrodes above and below the microwell with the target hybridoma. Results and discussion: Cell−based arrays were formed with the occupancy efficiency of over 90% in a few seconds by p-DEP. Antibodies secreted in microwell arrays were captured to discriminate target hybridomas in a few hours without repeated steps of a culture and a limiting serial-dilution. Furthermore, hybridomas trapped in microwells were retrieved from the cell−based array by applying an AC signal to band electrodes. The sequential system for forming cell−based arrays, discriminating hybridomas secreting specific antibodies, and retrieving target hybridomas was developed by using the novel microwell array device comprising 3-D microband array electrodes with an orthogonal arrangement.