An Online, Real-Time Microfluidic Biosensor for PFOA and PFOS

适用于 PFOA 和 PFOS 的在线实时微流体生物传感器

• 批准号: 10383822
• 负责人: Nicholas Csicsery
• 金额: $ 25.66万
• 依托单位: QUANTITATIVE BIOSCIENCES, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-12-01 至 2022-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10383822
• 关键词: Address; Agglutination; Agriculture; Binding; Biological Assay; Biosensing Techniques; Biosensor; Calibration; Detection; Engineering; Ensure; Environment; Escherichia coli; Fluorescence; Generations; Health; Heavy Metals; Home; Human; Image; Individual; Kinetics; Laboratories; Liver diseases; Malignant Neoplasms; Measures; Microfluidic Microchips; Microfluidics; Monitor; Nutrient; Poly-fluoroalkyl substances; Privatization; Process; Quality of life; Risk; Running; Safety; Signal Transduction; Small Business Innovation Research Grant; Specificity; Speed; Surface; Thyroid Diseases; Time; United States; Water; Water Supply; Work; base; commune; drinking; drinking water; epidemiology study; ground water; imaging platform; innovation; microbial; microfluidic technology; nanobodies; novel; novel strategies; operation; prototype; response; sensor; sensor technology; synthetic biology; vector; water testing

Access to clean, reliable water supplies is critical to our quality of life and our economy, and ensuring this access for generations to come will involve developing novel approaches to determining the safety and composition of potable water that are practical and affordable. Per- and polyfluoroalkyl substances (PFASs) are among the most ubiquitous and persistent contaminants plaguing groundwater in the United States, and human epidemiological studies have found associations between PFASs in drinking water and a number of adverse health conditions, from liver and thyroid disorders to various forms of cancer. The main objective of this SBIR proposal is to develop a customizable biosensor platform that uses engineered microbial sensor strains paired with microfluidic technology to continuously monitor water for PFASs. In order to demon- strate technical feasibility, QBI will perform these Specific Aims: Specific Aim 1: To identify and characterize the binding kinetics of nanobodies for PFOA and PFOS. For an engineered bacterial strain to be maximally effective as a sensor, it must be able to specifically and strongly bind its target in the environment. QBI will identify nanobodies that bind PFAS molecules and characterize their capture potential when surface displayed in an adsorbing E. coli strain. QBI will work with a local nanobody company, Abcore, to isolate a set of nanobodies that are enriched for specificity to the two targets and will then bring these nanobodies to their facility, clone them into their E. coli nanobody display vector, and screen and characterize them within their multiplexed microfluidic platform. Specific Aim 2: To develop a prototype for continuous and batch PFAS sensing. In order to use the newly developed sensor strains in a continuous monitoring platform, QBI will need to develop a novel as- say for measuring agglutination on a microfluidic-scale from many individual strain banks. QBI will begin by developing and optimizing an agglutination assay using the reduced set of surface-displayed nanobody strains, and then they will build upon previous results to transduce the agglutination signal to a fluores- cence response. Finally, QBI will optimize a microfluidic device to facilitate this assay and maximize the cellular fluorescence signal so that they can quantify the amount of contaminant present in the water. Successful completion of these Aims will serve to validate the use of nanobody-based sensing strains to achieve sensitive, selective, and continuous contaminant detection, making it of great utility to monitoring efforts aimed at tracking and assessing potential hazardous exposures. Beyond the ability to detect many different targets with a single on-line sensor, which is highly unique, the customizability and expandability of the platform using synthetic biology to engineer strains is transformative. This will enable QBI to continually expand their customer base as they continue to add sensing capabilities tailored to meet end-users' needs. 1

获得清洁、可靠的供水对我们的生活质量和经济至关重要， 未来几代人的这种访问将涉及开发新的方法来确定安全性， 和组成的饮用水是实用和负担得起的。全氟烷基和多氟烷基物质 PFASs是美国地下水中最普遍和持久的污染物之一， 美国和人类流行病学研究发现，饮用水中的PFAS与 一些不利的健康状况，从肝脏和甲状腺疾病到各种形式的癌症。的 该SBIR提案的主要目标是开发一种可定制的生物传感器平台， 传感器菌株与微生物技术相结合，可持续监测PFAS用水。为了恶魔化- 在技术可行性的前提下，QBI将实现以下具体目标： 具体目标1：确定和表征纳米抗体与PFOA和PFOS的结合动力学。 为了使工程菌株作为传感器最大限度地有效，它必须能够特异性地 在环境中牢牢地束缚住目标。QBI将识别结合PFAS分子的纳米抗体， 表征它们在吸附E中表面显示时的捕获电位。大肠杆菌菌株。QBI将发挥作用 与当地纳米抗体公司Abcore合作，分离出一组经过特定富集的纳米抗体， 两个目标，然后将这些纳米抗体带到他们的设施，克隆到他们的E。大肠杆菌纳米抗体 显示载体，并在其多路复用微电泳平台内筛选和表征它们。 具体目标2：开发用于连续和批量PFAS传感的原型。为了使用 新开发的传感器在连续监测平台中产生应变，QBI将需要开发一种新型的- 比如说用于在微生物学规模上测量来自许多个体菌株库的凝集。QBI将通过以下方式开始 开发和优化使用减少的表面展示纳米抗体组的凝集测定 菌株，然后他们将建立在以前的结果，以消除凝集信号的一个指标， 反应。最后，QBI将优化一种微电泳设备，以促进这种测定，并最大限度地提高 细胞荧光信号，这样他们就可以量化水中存在的污染物的数量。 这些目标的成功完成将有助于验证基于纳米抗体的传感菌株的使用，以实现 灵敏，选择性和连续的污染物检测，使其具有很大的效用，以监测工作，旨在 跟踪和评估潜在的危险暴露。除了能够探测到许多不同的目标， 一个高度独特的单一在线传感器，平台的可定制性和可扩展性 使用合成生物学来改造菌株是变革性的。这将使QBI能够不断扩大 他们的客户群，因为他们继续增加量身定制的传感能力，以满足最终用户的需求。 1