Robotic Whole-cell Patch Clamping Based on Three Dimensional Location for Adherent Cells

Whole-cell patch clamping is the gold-standard technology for electrical activities recordings of live single cells. To guarantee a high signal-to-noise ratio in recordings, a giga Omega-scale seal (gigaseal) between target cell membrane and the electrode needs to be formed first in whole-cell patch clamping. As lack of precise 3-D location methods for adherent cells, the operators have to roughly position the electrode to contact the target cell, easily resulting in gigaseal formation failures due to cell membrane penetration etc. For the first time, this paper introduced a fully automated whole-cell patch clamping system based on 3-D location for adherent cells under the bright field. First, a defocusing microscopy-based cell segmentation was proposed to locate adherent cell position in horizontal (X-Y) direction. Second, contact detection based on the measured electrode impedance was developed to measure the top surface position of adherent cell in vertical (Z) direction. Based on above work, an automated whole-cell patch clamping process was established. The experimental results on mouse osteoblasts showed that the system was able to successfully locate 94.7% (71/75) of cells with an average error of 0.3 μ m in horizontal direction. In vertical direction, the success rate and the average error of top surface position measurement for cells were 100% (20/20) and less than 4.65%, respectively. With precise 3-D positions of adherent cells, the success rate of whole-cell patch clamping using our method was significantly improved in comparison to manual operation results (70% Vs 40%). The proposed system in this paper provides an effective tool for electrical activities recordings of adherent cells in the future.