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.

测量血液中的蛋白质和核酸等生物分子（称为生物标志物）是检测癌症和其他疾病的一种有价值的方法，由多种蛋白质和核酸组成的样本显示出实现此目的的希望，但目前必须使用单独的时间进行测量。该项目使用一种称为 CRISPR（成簇规则间隔短回文重复序列）的生物结构来实现测量，CRISPR 可以被设计为结合核酸或用核酸标记的蛋白质并给出。仅当特定核酸或标签与 CRISPR 结合时，该方法才会被开发成一种快速、低成本的 3D 打印半自动化设备，以在同一测试中测量多种核酸和多种蛋白质生物标志物。还开发了一种称为电化学发光 (ECL) 的敏感电驱动光发射，作为一种更简单的检测方法，使用 3D 打印设备进行的血液测试将设计为在诊所或医生办公室内通过轻轻操作即可在短时间内进行。训练有素的技术人员可以在患者仍在办公室时以所谓的护理点模式快速周转，并可以在那时将其用作诊断的一部分。利用康涅狄格大学的项目支持研究生、本科生、科学教师和高中生的新 CRISPR 技术教育，活动包括通过 SECRET（探索化学研究经验和培训学院）举办 CRISPR 检测研讨会、发展早期大学。体验项目为中学生和科学教师提供便利，促进 CRISPR 阵列设计和制造方面的博士生培训，并为 UCONN 和 UCONN 的 NSF REU 项目以及通过 ACS SEED 项目的高中生提供研究机会。分子生物标志物在许多疾病的医学诊断中变得越来越有价值，与单一生物标志物相比，生物标志物组合显示出卓越的诊断效用，并且组合的蛋白质-miRNA 组合已显示出比单独蛋白质或 miRNA 显着改善的诊断预测。该项目将开发第一个使用 CRISPR 技术同时测量 miRNA 和蛋白质组合的分析阵列，新的 3D 打印阵列将能够分析 miRNA 蛋白质生物标志物面板，以进行广泛的疾病诊断。护理 (POC) 模式以及其他生物医学研究应用 除​​了荧光检测之外，还将利用 CDC 和 iPhone 相机检测来探索电化学发光 (ECL)，重点将放在高性能的 miRNA 和蛋白质生物标记物上。侵袭性前列腺癌的生物标记物将通过测量来自高级别前列腺癌患者和对照的 130 个现有人类血清样本中的目标 miRNA 和蛋白质来验证。该项目的智力优点包括使用新的工程策略的设计和开发。 CRISPR 分析技术可在 POC 中同时检测 miRNA 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。