Instrumentation Development for Parallel Measurement of an Ion Channel Array Chip

描述(由申请人提供)：用于药物发现和安全筛选的离子通道的电生理分析在高通量下执行是有问题的，因为离子通道必须结合到脂质双层膜中以能够测量它们的离子电导。因此，目前还没有用于离子通道筛选的高质量、高通量的分析方法。自动化膜片钳仪器的最新发展仍然比传统的可溶性蛋白质药物筛查的吞吐量低两个数量级以上，而且还需要昂贵的仪器、专门的细胞系和消耗品。对于现有的离子通道筛选方法，在信息质量、吞吐量和成本方面存在很大差距。Librede Inc.正在开发一种离子通道测量的替代技术，即在人造脂质双层膜中重建离子通道。Librede正在申请专利的无细胞人工双层配方具有显著提高产量和降低消耗品成本的潜力，同时需要更便宜的设备和训练有素的人员。Librede是由加州大学洛杉矶分校的这项技术的发明者创立的；我们现在正在努力将这项技术从学术实验室转移到商业上开发。在初步工作中，我们开发了廉价、一次性、可装运的双层阵列芯片，能够同时支持48个地点的离子通道测量。我们以前已经使用多路单通道放大器测量了这些芯片中的双层和离子通道；在这里提出的工作中，我们的目标是通过使用48通道膜片钳放大器同时测量芯片中的所有位置来展示更高的吞吐量。同时测量将是对我们技术的关键测试；我们将探索噪声、串扰和双层尺寸对我们同时测量整个芯片上的双层和结合离子通道的能力的影响。使用该仪器的系统获得的噪声和带宽将决定我们的信号检测和时间分辨率，并决定该平台用于离子通道高通量测量的可行性以及提高性能的途径。能够并行测量双层阵列中的离子通道将是这项技术走向商业化发展的一个重要里程碑，也是我们最终产品全面展示的初步步骤。我们将利用我们在第一阶段研究中获得的材料和经验进行第二阶段的开发，在这一阶段中，我们将把低成本、可批量生产的注塑可装运双层芯片与用于流体处理和平行离子通道测量的自动化仪器集成在一起。这将使我们更接近我们的目标，即通过消除药物筛选中的细胞培养和细胞操纵来降低离子通道筛选所需的成本和专业知识，从而显著提高产量和降低成本，使更有效地搜索离子通道药物成为可能。 公共卫生相关性：测量离子通道与药物的相互作用是药物发现和药物安全筛选的关键过程，但由于使用离子通道的困难，使用的现有过程缓慢、费力且昂贵。Librede最近开发了一种基于可运输的脂膜的离子通道测量平台，该平台成本低得多，使用起来也容易得多--这是世界上第一次。我们建议开发同时测量这些膜中包含的离子通道阵列的仪器。

DESCRIPTION (provided by applicant): Electrophysiological assays of ion channels for pharmaceutical discovery and safety screening are problematic to perform in high throughput because the ion channels must be incorporated into a lipid bilayer membrane to enable measurement of their ionic conductance. As a result, there are currently no high quality, high throughput assays for ion channel screening. Recent developments of automated patch clamp instrumentation are still over two orders of magnitude lower throughput than conventional drug screening for soluble proteins and also require expensive instrumentation, specialized cell lines, and consumables. For existing methods of ion channel screening, there is a large gap in information quality, throughput, and cost. Librede Inc. is developing an alternative technology for ion channel measurement in which the ion channels are reconstituted in artificial lipid bilayer membranes. Librede's patent pending formulation of cell-free artificial bilayers has the potential for significantly higher throughput and lower consumable costs, while requiring less expensive equipment and trained personnel. Librede was founded by the UCLA inventors of this technology; we are now working to transfer this technology from the academic laboratory and develop it commercially. In preliminary work, we have developed inexpensive, disposable, shippable bilayer array chips capable of supporting ion channel measurement in 48 sites simultaneously. We have measured bilayers and ion channels in these chips previously using a multiplexed single channel amplifier; in the work proposed here, we aim to demonstrate higher throughput by measuring all sites in the chip simultaneously using a 48 channel patch clamp amplifier. Simultaneous measurement will be a key test of our technology; we will explore the effects of noise, crosstalk, and bilayer size on our ability to measure bilayers and incorporated ion channels over the entire chip at once. The noise and bandwidth achieved with our system using this instrument will determine our signal detection and temporal resolution and determine the feasibility of this platform for high throughput measurement of ion channels as well as paths to improvements in performance. The ability to measure ion channels in bilayer arrays in parallel will be a major milestone in the development of this technology toward commercialization, and is a preliminary step of the full demonstration of our final product. We will use the materials and experience gained in our Phase I research for Phase II development in which we will integrate low cost mass-producible injection molded shippable bilayer chips with automatable instrumentation for fluid handling and parallel ion channel measurement. This will bring us closer to our goal of reducing the cost and expertise required for ion channel screening by eliminating cell culture and cellular manipulation in pharmaceutical screening, thus significantly increasing throughput and decreasing costs, enabling more effective searches for ion channel drugs. PUBLIC HEALTH RELEVANCE: Measurement of ion channel interactions with drugs is a key process in drug discovery and drug safety screening, but due to the difficulty in working with ion channels the existing processes used are slow, laborious, and expensive. Librede has recently developed a platform for ion channel measurement which is much less expensive and much easier to use, based on lipid membranes that can be shipped-a world first. We propose here to develop instrumentation for simultaneously measuring arrays of ion channels contained in these membranes.