Development of a prototype system for assaying exocytosis from individual cells

描述（由申请人提供）：细胞通过胞吐分泌激素和神经递质。了解胞吐是如何被调节的，以及药物和毒素是如何抑制胞吐的，具有广泛的医学意义。SBIR项目的总体目标是使用微加工技术开发一种强大且易于使用的原型设备，该设备可以比现有方法快10- 100倍并便宜地分析单个细胞释放的递质，从而大大加快生物医学研究的步伐。此外，该设备将提供全新的功能，例如能够同时成像荧光标记的囊泡，同时测量从同一囊泡释放的透明电化学电极。具体目的是开发和测试透明多孔电化学电极阵列装置的原型。每个井将是一个独立的实验室，底部将包含一系列电化学电极。单个细胞将通过使用细胞大小的微孔和细胞粘附分子的模式连接到每个电极上。电极将是透明的，并在薄玻璃基板上形成图案，以允许同时进行高分辨率光学和电化学测量。电极阵列将使用专有方法进行多路复用，以简化与外部放大器所需的连接数量。第一阶段的具体里程碑是使用单个多孔设备在一小时内记录至少100个细胞的定量胞吐。实现这一里程碑将证明，原型设备获取数据的速度比目前使用碳纤维微电极的方法快10倍，因此将确定在实际平台上可以实现吞吐量的根本改进。该设备的潜在市场包括生物医学研究界、针对胞吐作用进行高通量药物筛选的制药行业、不使用实验动物进行肉毒中毒自动检测的临床市场，以及使用基于细胞的生物传感器检测神经毒素生物武器的国防/国土安全市场。Exocytronics LLC将咨询潜在的学术用户/客户，为设备设计提供反馈，将从经验丰富的企业合作伙伴那里获得营销建议，并将在MU生命科学企业孵化器中获得创业支持。

DESCRIPTION (provided by applicant): Cells secrete hormones and neurotransmitter via exocytosis. Understanding how exocytosis is regulated and how pharmaceuticals and toxins inhibit exocytosis is of broad medical significance. The overall goal of this SBIR project is to use microfabrication technology to develop a robust and easy-to-use prototype device that can assay release of transmitter from individual cells 10- 100 fold faster and cheaper than current approaches and thereby greatly accelerate the pace of biomedical research. In addition, the devices will provide entirely novel capabilities such as the ability to simultaneously image a fluorescently labeled vesicle while measuring release from the same vesicle with an underlying transparent electrochemical electrode. The Specific Aim is to develop and test a prototype transparent multi-well electrochemical-electrode array device. Each of the wells will be a separate experimental chamber and will contain an array of electrochemical electrodes on the bottom. Individual cells will be docked to each electrode through the use of cell-sized microwells and patterning of cell-adhesion molecules. The electrodes will be transparent and patterned on a thin glass substrate to allow simultaneous high-resolution optical and electrochemical measurements. The electrode arrays will be multiplexed using a proprietary approach to simplify the number of connections required to an external amplifier. The specific milestone for Phase 1 is to record quantal exocytosis from at least 100 cells within one hour using a single multi-well device. Achieving this milestone will demonstrate that the prototype device can acquire data ~10-fold faster than the current method using carbon-fiber microelectrodes, and thus will establish that radical improvements in throughput can be achieved in a practical platform. Potential markets for the devices include the biomedical research community, the pharmaceutical industry to carry out high-throughput screening of drugs that target exocytosis, a clinical market to perform automated assays of botulism without the use of laboratory animals and a defense / homeland security market to detect neurotoxin bioweapons with cell-based biosensors. Exocytronics LLC will consult potential academic users / customers to provide feedback on device design, will obtain marketing advice from an experienced corporate partner and will obtain entrepreneurial support while housed in the MU Life Sciences Business Incubator.