A Miniaturized Fluidic Chip For Long-term Maintenance and Impedance Monitoring of

描述（由申请人提供）：该SBIR项目旨在通过延长培养的哺乳动物细胞的活力来开发小型化的基于细胞的生物传感器，以实现低成本、便携性和延长的保质期。我们将通过创建尺寸比目前可用的电电池基板阻抗传感（ECIS）系统小约100倍的微芯片来展示我们当前技术的优势。由于更小的芯片尺寸，将大大降低成本和介质存储和更换量。我们还将通过更换为适当的CO2非依赖性缓冲系统来优化培养基条件，以延长所选哺乳动物细胞系在环境条件下至少45天的活力。此外，我们将结合多参数细胞传感，以改善毒性检测，而不是使用单一的细胞阻抗参数。这种小的，低成本的系统具有延长的保质期将具有显着的价值，作为一个便携式现场设备筛选毒物和病原体在临床环境中，食品和供水，以及在高通量应用中的目标验证制药工业。该SBIR提案的具体目标包括：1）设计和制造用于阻抗测量的小型化电路，2）设计和制造不透CO2的微流体卡，3）在不依赖CO2的培养基中培养细胞并监测在微流体环境中45天期间和之后的阻抗变化，和4）用至少三种已知的化学毒物评价所开发的微流体卡的毒物响应性，并与市售ECIS系统进行比较。 公共卫生相关性：基于细胞的生物传感器（Cell-based biosensors，CBB）在检测病原体、毒素或生物活性化合物方面具有巨大的潜力，可用于临床、制药、环境、生物安全和高通量筛选（High-throughput screening，HTS）等领域。在筛选食品和农产品时尽早使用这些标准将是有益的，因为食品、水和农业的安全对公共卫生和经济有重大影响，因而是最令人关切的问题。哺乳动物细胞为基础的生物传感器正在开发中，在诊所或医院现场使用，作为生物防御应用和环境监测的第一反应。目前，这些基于哺乳动物细胞的系统是昂贵的，因为它们的保质期短，因为哺乳动物细胞在没有大的外部培养基递送和维持系统的情况下最难以保持培养数天。低保存期和高维护成本限制了在现场操作和HTP筛选应用中使用其他高效筛选工具。因此，需要取得重大进展来改进目前的系统。微制造、微流体和细胞培养的集成有能力通过制造小型、便携且具有成本效益的传感器来改变当前的情况，因为制造它们所需的原材料较少。此外，小尺寸降低了培养基和培养物的要求。此外，优化实地使用的介质条件将使用户能够使用这些介质。

DESCRIPTION (provided by applicant): This SBIR project is designed to develop miniaturized cell-based biosensors for low cost, portability and enhanced shelf life by extending the viability of the cultured mammalian cells. We will demonstrate the benefit of our current technology by creating micro-chips with approximately 100 times smaller dimensions than the currently available Electrical Cell- Substrate Impedance Sensing (ECIS) systems. Because of the smaller chip dimensions there will be significant reduction in cost and media storage and replacement volumes. We will also optimize the media conditions by changing to appropriate CO2- independent buffer systems to extend the viability of the chosen mammalian cell line for a period of at least 45 days under ambient conditions. Moreover, we will incorporate multiparameter cellular sensing for improved toxicity detection over using a single parameter of cellular impedance. This small, low cost system with extended shelf life will have significant value as a portable field device for screening toxicants and pathogens in clinical settings, food and water supplies, as well as in high throughput applications for target validation in pharmaceutical industry. Specific aims of this SBIR proposal include: 1) Design and manufacture miniaturized circuits for impedance measurements, 2) Design and manufacture CO2- impervious microfluidic cards, 3) Propagate cells in CO2- independent media and monitor change in impedance during and after 45 days in a microfluidic environment, and 4) Evaluate toxicant responsiveness of the developed microfluidic card with at least three known chemical toxicants and compare with commercially available ECIS systems. PUBLIC HEALTH RELEVANCE: Cell-based biosensors (CBBs) have tremendous potential to be used as efficient, reliable and sensitive tools for the detection of pathogens, toxins or bioactive compounds for clinical, pharmaceutical, environmental, biosecurity and high-throughput screening (HTS) applications. Their early use in the screening of food and agriculture products would be beneficial as the safety of food, water and agriculture is of ultimate concern because of its significant impact on public health and economy. Mammalian cell-based biosensors are being developed for on-site use in clinics or hospitals as a first response in biodefense applications and for environmental monitoring. Currently these mammalian cell based systems are expensive because of their short shelf life, as mammalian cells are most difficult to keep in culture for several days without large external media delivery and maintenance systems. The low shelf life and high maintenance cost limits the use of an otherwise highly effective screening tool in field operations and in HTP screening applications. Therefore, significant advances are required to improve the present systems. The integration of microfabrication, microfluidics and cell culture have the ability to change the current scenario by fabricating sensors that are small, portable and cost effective because less raw material is required to fabricate them. In addition, the small size reduces the media and culture requirements. Moreover, the optimization of media conditions for field use would make them accessible to the user.