Highly Integrated Nucleic-Acid Analysis Using Graphene Bioelectronics

使用石墨烯生物电子学进行高度集成的核酸分析

• 批准号: 10584520
• 负责人: Jinglei Ping
• 金额: $ 18.99万
• 依托单位: UNIVERSITY OF MASSACHUSETTS AMHERST
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-01 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10584520
• 关键词: Affinity; Animal Model; Biological Assay; Biological Markers; Biopsy; Biopsy Specimen; Blinded; Buffers; Clinical; Clinical Trials; Complex; Consumption; Core Facility; DNA; DNA Probes; Development; Devices; Diagnosis; Diagnostic; Disease; Disease Progression; Electrodes; Electronics; Generations; Genomics; Goals; Hospitals; Hour; Immunoassay; Intervention; Investigation; Measurable; Measures; MicroRNAs; Monitor; Mus; Noninfiltrating Intraductal Carcinoma; Nucleic Acids; Oligonucleotides; Operative Surgical Procedures; Outcome; Pathway interactions; Patients; Performance; Plasma; Population; Prognosis; Progressive Disease; Property; Public Health; Reaction; Recurrence; Research; Risk Assessment; Sampling; Signal Pathway; Signal Transduction; Technology; Testing; Therapeutic; Time; Transducers; Transistors; Translating; Translational Research; Validation; Visit; Xenograft Model; bioelectronics; breast cancer progression; circulating microRNA; clinical care; clinical translation; cohort; cost; detection limit; diagnostic technologies; disease diagnosis; early screening; genomic predictors; graphene; human subject; improved; individualized medicine; liquid biopsy; malignant breast neoplasm; micro-total analysis system; mouse model; new therapeutic target; next generation; outcome prediction; overtreatment; pH gradient; point of care; point-of-care diagnosis; pre-clinical; prevent; prognostic; programs; response; screening; sensor; targeted treatment; technology platform; technology validation; treatment response; treatment strategy; user-friendly; voltage

PROJECT SUMMARY The circulating population of microRNAs in biofluids are ideal biomarkers for various diseases. Point-of-care profiling of circulating microRNAs is in insatiable demand, but typical approaches, e.g., immunoassays and microRNA assays are lab-based/centralized, expensive ($400–1,000/test), and time-consuming (>6 hours). We will develop a highly integrated, all-nanobioelectronic platform technology for multiplex, high-accuracy circulating-microRNA analysis that is capable of profiling circulating microRNAs in a 50-μL plasma with ultra- high sensitivity (sub-fM) and efficiencies in time (<40 minutes) and cost (<$10/test), thereby enabling high- performance circulating-microRNA analysis at the point of test. The novelty of the program is to harness graphene-based bioelectronics to integrate circulating microRNA isolation, concentration, amplification, and quantification into a self-contained device. In order to proof the concept of this technology, the program will include the development and validation of two generations of graphene-based analytical platforms, GAP1 and GAP2. Three specific aims with measurable milestones will be pursued. (1) We will demonstrate that multiple microRNA analytes can be amplified via hybridization chain reaction on a probe-functionalized graphene sensor array and the analyte concentrations can be readily interrogated by the graphene sensor array and translated into electrical signals. We will develop GAP1 to selectively quantify eight pre-selected target microRNAs (MDCIS8) spiked in 5-μL buffer. The detection limit of the specific microRNAs is expected to be at fM level. (2) We will demonstrate that target circulating microRNAs can be isolated from plasma by immobilizing them on a DNA-functionalized graphene electrode and releasing them into a small-volume simple cargo solution upon the generation of pH gradient by applying voltage bias between the graphene-DNA electrode with a bare graphene electrode. We will develop a graphene-based circulating-microRNA isolation module, combine the module with GAP1 to form GAP2, and use GAP2 to profile circulating MDCIS8 in lysed samples of 50-μL plasma from NSG mice. The GAP2 is expected to concentrate the microRNAs by >5× and deliver sub-fM level sensitivity. (3) We will demonstrate the feasibility of using this platform technology for diagnostic applications. We will use GAP2 to quantify circulating MDCIS8, whose expression levels are indicative to pre-invasive breast cancer, in 50-μL plasma samples from a user blinded cohort of the MIND murine model. The profiling result will be analyzed to predict the progression of pre-invasive breast cancer whose rapid, inexpensive diagnosis remains a challenge. The GAP2 prediction outcome will be combined with that based on surgical biopsy to establish the accuracy of the technology for progression prediction. The expected prediction accuracy is >96%. If successful, the technology will offer a new pathway to next-generation point-of-care genomic diagnostic/prognostic micro total analysis systems that would be cheap enough and user friendly enough to be used in various clinical settings.

项目摘要 生物流体中的microRNA循环种群是各种疾病的理想生物标志物。护理点 循环microRNA的分析是无法满足的需求，但典型的方法，例如免疫测定和 MicroRNA测定基于实验室/集中式，昂贵（400-1,000美元/测试）和耗时（> 6小时）。我们 将开发一种高度集成的全纳米电代理平台技术，用于多重，高临界性 循环中的微生纳分析能够分析50μl血浆中循环microRNA， 高灵敏度（sub-fm）和效率的时间（<40分钟）和成本（<$ 10/测试），从而使高 测试点的性能循环 - 微洋分析。该计划的新颖性是要利用 基于石墨烯的生物电子学以整合循环的microRNA隔离，浓度，扩增和 数量成一个独立设备。为了证明这项技术的概念，该计划将 包括两代基于石墨烯的分析平台的开发和验证，GAP1和 GAP2。将实现三个特定目标，以可衡量的里程碑。 （1）我们将证明这是多个 MicroRNA分析物可以通过探针功能化石墨烯传感器上的杂交链反应扩增 阵列和分析物浓度很容易被石墨烯传感器阵列询问并翻译 进入电信号。我们将开发GAP1以选择性量化八个预选的目标microRNA （MDCIS8）在5μl缓冲液中升高。特定microRNA的检测极限预计在FM水平上。 （2） 我们将证明，可以通过将它们固定在A上来隔离等离子体。 DNA官能化石墨烯电极，并将其释放到小体积的简单货物溶液中 通过用裸石石墨烯在石墨烯-DNA电极之间施加电压偏置来产生pH梯度 电极。我们将开发一个基于石墨烯的循环丝菌隔离模块，将模块与 GAP1形成GAP2，并使用GAP2在NSG的50μl等离子体的裂解样品中介绍循环的MDCIS8 老鼠。 GAP2预计将以> 5倍的浓度将microRNA浓缩，并提供次-FM水平敏感性。 （3）我们 将证明将此平台技术用于诊断应用的可行性。我们将使用GAP2来 量化循环的mDCIS8，其表达水平在50μl中表示为侵入性乳腺癌 来自用户盲鼠模型的用户盲人队列的等离子体样本。分析结果将进行分析 预测侵入性乳腺癌的进展，其快速，廉价的诊断仍然是一个挑战。 GAP2预测结果将基于手术活检，以确定的准确性 进展预测的技术。预期的预测准确性> 96％。如果成功， 技术将为下一代医疗点基因组诊断/预后微型总计提供新的途径 分析系统足够便宜且用户友好，足以在各种临床环境中使用。