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

描述(申请人提供)：该SBIR项目旨在开发基于细胞的微型生物传感器，通过延长培养的哺乳动物细胞的活力来实现低成本、便携性和延长保质期。我们将通过制造尺寸比当前可用的电池板-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。由于芯片尺寸较小，成本以及媒体存储和更换数量将大幅减少。我们还将通过改用适当的不依赖二氧化碳的缓冲系统来优化培养条件，以延长所选哺乳动物细胞系在环境条件下的活力至少45天。此外，我们将结合多参数细胞传感，以改进毒性检测，而不是使用单一参数的细胞阻抗。这种体积小、成本低、保质期长的系统将作为一种便携式现场设备，在临床环境、食品和水供应中筛选毒物和病原体，以及在制药行业的目标验证的高通量应用中具有重要价值。这项SBIR建议的具体目标包括：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)在不依赖二氧化碳的介质中繁殖细胞，监测微流体环境中45天内和45天后的阻抗变化，以及4)评估开发的微流体卡对至少三种已知化学毒物的毒物响应性，并与商用ECIS系统进行比较。 与公共卫生相关：基于细胞的生物传感器(CBB)具有巨大的潜力，可以作为有效、可靠和灵敏的工具来检测病原体、毒素或生物活性化合物，用于临床、制药、环境、生物安全和高通量筛选(HTS)应用。及早在食品和农产品筛查中使用它们将是有益的，因为食品、水和农业的安全是最终令人关切的问题，因为它对公众健康和经济有重大影响。以哺乳动物细胞为基础的生物传感器正在开发中，用于诊所或医院的现场使用，作为生物防御应用和环境监测的第一反应。目前，这些以哺乳动物细胞为基础的系统价格昂贵，因为它们的保质期很短，因为如果没有大型的外部介质输送和维护系统，哺乳动物细胞很难在培养中保持几天。低保质期和高维护成本限制了在外地业务和HTP筛查应用中使用原本非常有效的筛查工具。因此，需要取得重大进展，以改进目前的制度。微制造、微流体和细胞培养的集成能够通过制造小型、便携和成本效益高的传感器来改变目前的情况，因为制造这些传感器所需的原材料更少。此外，较小的尺寸降低了对媒体和文化的要求。此外，优化现场使用的媒体条件将使用户能够接触到它们。

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.