Microfluidic-based high-efficiency cell fusion for studying nuclear reprogramming

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DESCRIPTION (provided by applicant): We propose to develop a new microfluidic technology to efficiently create 1,000's of fusion events between embryonic stem cells and somatic cells in order to study early events in nuclear reprogramming. Our device uses a three-step loading procedure and a special geometry to efficiently arrange cells side-by-side to create a large fraction of properly paired hybrid cells. We are developing this technology to study nuclear reprogramming, which is the process whereby a somatic cell's nuclear program is rebooted to bring that cell back into a more primitive, pluripotent state. Factors within pluripotent or totipotent cells can reprogram somatic cells, and the current approaches center on methods of transferring these factors from one cell to the other. The standard approach to reprogramming nuclear transfer, in which the somatic cell nucleus is transplanted into an unfertilized egg is tedious and limited to small numbers of cells. Cell fusion, whereby a somatic cell is fused to an embryonic stem cell, merging the cellular contents, can also be used to reprogram the somatic cell's nucleus. Cell fusion has the potential to be easier and more scalable than nuclear transfer. One significant technical limitation that prevents the widespread use of this approach for studying reprogramming is that current approaches to cell fusion use randomly paired cells. This results in a small number of desired hybrid fusions (stem cell + somatic cell) in a large background of unwanted fusions and unfused cells. In order to purify the desired fusions from this background, researchers undertake antibiotic selection steps that add days to the procedure and prevent researchers from studying early events in nuclear reprogramming. By developing a device to efficiently pair cells, we are addressing the dominant technological challenge in the creation of hybrids, and believe that our device will have significant impact for studying fusion-induced reprogramming.