A Cortisol Sensing Enzyme System: A New Platform Utilizing Dehydrogenases in Biosensors

项目摘要 第一阶段SBIR的目标是开发一种新的平台，使酶能够用作酶 在电流生物传感器中。无论平台或测量模态如何，所有生物传感器设计都需要 使用酶来正常发挥作用。生物传感器是测量化合物的装置， 人体健康FDA已经批准用于监测葡萄糖的电流生物传感器， 目前，一些新的生物传感器平台正在开发中。生物传感器需要酶 其对于要测量的化合物是特异性的。 该提案将通过开发一种酶系统来展示我们平台的实用性， 在护理点设备和生物传感器中监测皮质醇。发展的主要障碍是 连续皮质醇监测系统是缺乏一种酶，可以作为设备制造的一部分。 此时，还没有描述适合于皮质醇检测和适合于生物传感应用的酶 或商业化。这说明了开发新的分析物特异性酶系统的重要性 对人类健康很重要。一旦概念验证在第一阶段得到证明，我们的第二阶段将扩展 用于与其他酶一起使用以监测对人类健康和疾病重要的分析物的平台。相 II类靶点包括马来酸脱氢酶、GABA脱氢酶、琥珀酸脱氢酶、丙酮酸脱氢酶 脱氢酶、乙醛脱氢酶和醇脱氢酶。这些脱氢酶 或者没有已知的氧化酶对应物，或者具有不稳定的氧化酶对应物， 酶基本上无用的生物传感器制造的研究在自由移动的动物。阶段结束时 II SBIR，我们将有一个优化的平台，生产高度稳定的脱氢酶，用于 生物传感器此外，还将开发适合人类使用的功能性皮质醇生物传感器。没有 目前存在连续测量体内皮质醇水平的诊断装置。 在这项研究中，我们将使用我们提出的平台，通过将两个新的 酶产生的信号可以在电极上以高灵敏度检测。这种双重酶 这种以共价连接的辅酶分子为特征的方法是高度创新的， 第一次连续监测对人体健康重要的分析物，如皮质醇。基于新设备 这项技术将使临床医生能够监测和管理疼痛，压力和其他负面的条件， 影响人类健康。我们的平台将对其他设备的开发和制造产生积极影响 用于新颖的传感应用。第一阶段的交付将是一个耦合酶系统的原型 展示了监测皮质醇水平的选择性、灵敏性和有效机制的概念验证， 组织和体液。成功完成这项提案是建立一个 一种连续监测所有高危人群皮质醇的方法。

Project Abstract The goal of this Phase I SBIR is to develop a new platform that enables dehydrogenases to be used as enzymes in amperometric biosensors. Regardless of the platform or measurement modality, all biosensor designs require the use of an enzyme to properly function. Biosensors are devices that measure compounds important for human health. FDA approval has already been achieved for amperometric biosensors that monitor glucose in humans, and several new biosensor platforms are currently under development. Biosensors require an enzyme that is specific for the compound to be measured. This proposal will showcase our platform’s utility by developing an enzyme system that provides the ability to monitor cortisol in point-of-care devices and biosensors. The major impediment to the development of a continuous cortisol monitoring system is the lack of an enzyme that can be used as part of device fabrication. At this time, no enzyme suitable for cortisol detection and suitable for biosensing applications has been described or commercialized. This illustrates the importance of developing new enzyme systems specific for analytes important in human health. Once proof-of-concept is demonstrated in Phase I, our Phase II will extend the platform for use with other dehydrogenases to monitor analytes important in human health and disease. Phase II targets include maleate dehydrogenase, GABA dehydrogenase, succinate dehydrogenase, pyruvate dehydrogenase, acetaldehyde dehydrogenase, and alcohol dehydrogenase. These dehydrogenase enzymes either have no known oxidase counterpart or have oxidase counterparts that are so unstable as to render the enzyme essentially useless in biosensor fabrication for studies in freely moving animals. At the end of the Phase II SBIR, we will have an optimized platform that produces highly stabilized dehydrogenase enzymes for use in biosensors. In addition, a functioning cortisol biosensor suitable for use in humans will be developed. No diagnostic device presently exists to continuously measure cortisol level in vivo. In this study we will use our proposed platform to create a cortisol enzyme system by coupling together two novel enzymes to produce a signal that can be detected on an electrode with high sensitivity. This dual enzyme approach that features a covalently attached co-enzyme molecule, is highly innovative and will enable, for the first time, the continuous monitoring analytes important to human health such as cortisol. New devices based on this technology will allow clinicians to monitor and manage pain, stress, and other conditions that negatively impact human health. Our platform will positively impact the development and manufacturing of other devices for novel sensing applications. The Phase I deliverable will be a prototype of the coupled enzyme system demonstrating proof-of-concept for a selective, sensitive and efficient mechanism of monitoring cortisol levels in tissues and fluids. The successful completion of this proposal is the necessary first step to establishing a continuous method of monitoring cortisol in all at risk individuals.