CRISPR Microfluidic Array for Simultaneous Multiplexed Sensing of Proteins and miRNAs

用于同时多重检测蛋白质和 miRNA 的 CRISPR 微流体阵列

• 批准号: 2231490
• 负责人: James Rusling
• 金额: $ 49.96万
• 依托单位: University of Connecticut
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-01 至 2026-04-30
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2231490&HistoricalAwards=false
• 关键词: CRISPR; Microfluidic; Array; Simultaneous; Multiplexed

Measurement of biomolecules like proteins and nucleic acids, called biomarkers, in blood, is a valuable approach to detect cancers and other diseases. Samples consisting of several proteins and nucleic acids show promise for this purpose, but at present must be measured using separate, time-consuming methods. This project uses a biological construct called CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) to enable measurements. CRISPR can be designed to bind a nucleic acid or a protein labeled with a nucleic acid and give a signal only when that specific nucleic acid or label binds to CRISPR. This approach will be developed into a fast, low-cost 3D-printed, semi-automated device to measure several nucleic acids and several protein biomarkers in the same test. The project will also develop sensitive electrically-driven light emission called electrochemiluminescence (ECL) as a simpler detection method. The blood test using the 3D-printed device will be designed to run in a short time in a clinic or physician’s office by a lightly trained technician. This will allow fast turnaround in what is called point of care mode. Results will be available to the physician while the patient is still in his or her office and can be used at that time as part of the diagnosis. This project will utilize programs at Univ. of Connecticut to support education in new CRISPR technology for graduate students, undergraduates, science teachers and high school students. Activities include holding workshops on CRISPR detection through SECRET (School of Exploratory Chemistry Research Experience and Training), developing Early College Experience programs for secondary school students and science teachers, facilitating training of Ph.D. students in CRISPR array design and fabrication, and providing research opportunities for undergraduates at UCONN and in UCONN’s NSF REU program, as well as high school students through the ACS SEED program.Molecular biomarkers have become increasingly valuable in medical diagnostics for many diseases. Panels of biomarkers, as opposed to single biomarkers, show superior diagnostic utility and combined protein-miRNA panels have shown significantly improved diagnostic predictions over separate protein or miRNA panels. This project will develop the first analytical array using CRISPR technology to simultaneously measure combinations of miRNAs and proteins. The new 3D-printed array will be able to analyze miRNA-protein biomarker panels for a wide range of disease diagnostics in point-of-care (POC) mode, as well as in other biomedical research applications. In addition to fluorescent detection, electrochemiluminescence (ECL) will be explored with CDC and iPhone camera detection. The focus will be on miRNA and protein biomarkers that are high performing biomarkers for aggressive prostate cancer. Assays will be validated by measuring target miRNAs and proteins in 130 existing human serum samples from patients with high grade prostate cancers and controls. The intellectual merit of this project involves design and development of the first engineering strategy using a new CRISPR assay technology to detect panels of miRNAs and protein simultaneously at POC.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

测量血液中的生物分子，如蛋白质和核酸，称为生物标志物，是检测癌症和其他疾病的有价值的方法。由几种蛋白质和核酸组成的样品显示出用于此目的的前景，但目前必须使用单独的，耗时的方法进行测量。该项目使用一种名为CRISPR（重复的规则间隔短回文重复序列）的生物结构来进行测量。CRISPR可以被设计为结合核酸或用核酸标记的蛋白质，并且仅当该特定核酸或标记物结合CRISPR时才给出信号。这种方法将被开发成一种快速、低成本的3D打印半自动化设备，用于在同一测试中测量多种核酸和多种蛋白质生物标志物。该项目还将开发称为电化学发光（ECL）的灵敏电驱动光发射，作为一种更简单的检测方法。使用3D打印设备的血液测试将被设计为在诊所或医生办公室由受过轻度训练的技术人员在短时间内运行。这将允许所谓的护理点模式的快速周转。结果将提供给医生，而病人仍然在他或她的办公室，并可以在当时作为诊断的一部分。该项目将利用康涅狄格大学的项目，为研究生、本科生、科学教师和高中生提供CRISPR新技术教育。 活动包括通过SECRET（探索性化学研究经验和培训学校）举办CRISPR检测研讨会，为中学生和科学教师开发早期大学体验计划，促进博士培训。在CRISPR阵列设计和制造的学生，并提供研究机会，为本科生在康州大学和康州大学的NSF REU计划，以及高中学生通过ACS种子计划。分子生物标志物已成为越来越有价值的医学诊断许多疾病。与单一生物标志物相反，生物标志物组显示出上级诊断效用，并且组合的蛋白质-miRNA组显示出比单独的蛋白质或miRNA组显著改善的诊断预测。该项目将开发第一个使用CRISPR技术的分析阵列，以同时测量miRNA和蛋白质的组合。新的3D打印阵列将能够分析miRNA-蛋白质生物标志物面板，用于护理点（POC）模式下的广泛疾病诊断以及其他生物医学研究应用。除了荧光检测外，还将使用CDC和iPhone相机检测探索电化学发光（ECL）。 重点将放在miRNA和蛋白质生物标志物上，它们是侵袭性前列腺癌的高性能生物标志物。将通过测量来自高级别前列腺癌患者和对照组的130个现有人类血清样本中的靶miRNA和蛋白质来验证分析。该项目的智力价值包括设计和开发第一个工程策略，使用新的CRISPR检测技术在POC中同时检测miRNAs和蛋白质组。该奖项反映了NSF的法定使命，并通过使用基金会的智力价值和更广泛的影响审查标准进行评估，被认为值得支持。