UNS: Role of Reporter and Membrane Properties for the Sensitivity of Viral Nanoparticle Lateral Flow Assays

1511789康拉德，杰辛塔研究人员已经证明，通过使用经过改造的生物活性纳米颗粒可以显著提高灵敏度。拟议的方法开发的应用，如果成功，将简化包括食品、水和医学样本在内的各种环境中的病原体检测。拟议的研究的目标是识别、理解并最终设计在侧向流动分析(LFA)中提供灵敏反应的噬菌体报告器和膜的特性。假设是调整噬菌体纳米颗粒的性质，LFA膜将增强噬菌体的运输和结合，从而降低LFA的检测下限。其具体目的是：(1)改变噬菌体形态以了解作为LFA报告分子的噬菌体性能；(2)确定增强噬菌体转运和粘附膜的膜特性；以及(3)通过定向进化选择性能优越的噬菌体。当不同形状和大小的噬菌体通过LFA膜扩散并结合到LFA膜上时，将用共聚焦显微镜成像。利用高通量的图像处理算法，噬菌体的定向和分散，结合将与膜的性质和孔大小相关。定向进化将被用于从数十亿个候选文库中选择具有更好的运输和特异性的噬菌体。智力优势：该项目将PI在成像和受限传输方面的专业知识与co-PI在色谱相互作用和噬菌体工程方面的专业知识相结合，以开发灵敏度更高的LFA。噬菌体记者作为LFA记者具有潜在的变革性，能够以极低的非特异性结合实现快速、廉价和超灵敏的检测。这项拟议的研究将建立超灵敏的LFA作为一种广泛适用的平台技术。非球形纳米粒子在多孔介质中的输运过程还没有得到充分的研究，本文将利用噬菌体作为一种新的模型体系来研究复杂受限介质中的分散问题。最后，这项建议引入了噬菌体进化与分离，作为一种新的方法来设计噬菌体，以应用于新兴的纳米生物技术领域。更广泛的影响：病毒纳米颗粒LFA技术可以很容易地与微流体和基于智能手机的荧光成像相结合，以产生一种廉价但超灵敏的使用点诊断工具。生物分离在现代生物制药制造的成本和工艺复杂性中占据主导地位；不需要训练有素的调查人员的过程分析(PAT)技术将立即影响这一领域。此外，一种可现场携带的分析方法可能被证明对环境污染、食品安全监测和医疗诊断有用。休斯顿大学是美国种族多样性排名第二的研究型大学，被指定为为拉美裔服务的机构，为扩大代表不足群体的参与提供了肥沃的招聘土壤。PIS将继续进行针对当地科学界的外联工作，通过为学术和工业科学家以及整个社会组织德克萨斯州软物质会议，通过为学术会议撰写和呈现科普广播节目，以及为我们的独创性引擎。

CBET - 1511789 Conrad, Jacinta The investigators have shown that significantly improved sensitivity can be achieved by using engineered bio-active nanoparticles. The applications of the proposed method development, if successful, will simplify pathogen detection in a variety of contexts, including food, water and medical samples.The objective of the proposed research is to identify, understand, and ultimately engineer the properties of phage reporters and membranes that provide sensitive response in lateral flow assays (LFA). The hypothesis is tailoring the properties of phage nanoparticles and LFA membranes will enhance phage transport and binding and as a result reduce LFA limit of detection. The specific aims are to: (1) vary phage morphology to understand phage performance as LFA reporters; (2) identify membrane properties that enhance phage transport and adhesion onto the membrane; and (3) select phage for superior performance using directed evolution. Phage reporters of varied shape and size will be imaged with confocal microscopy as they diffuse through and bind to LFA membranes. Using high-throughput image-processing algorithms, phage orientation and dispersion, binding will be correlated to membrane properties and pore size. Directed evolution will be used to select phage with improved transport and specificity from libraries of billions of candidates. Intellectual Merit: This project integrates the expertise of the PI in imaging and confined transport with that of the co-PI in chromatographic interactions and phage engineering to develop LFAs of increased sensitivity. Phage reporters are potentially transformative as LFA reporters, enabling fast, inexpensive and ultrasensitive detection with extremely low non-specific binding. The proposed research will establish ultrasensitive LFAs as a broadly-applicable platform technology. The transport of nonspherical nanoparticles in porous media is under-investigated, and the proposed work will use phage as a new model system in for studing dispersion in complex and confined media. Finally, this proposal introduces phage evolution with separation as a novel method to engineer bacteriophage for applications in the emerging area of nanobiotechnology. Broader Impacts: Viral nanoparticle LFA technology can be readily integrated with microfluidics and smartphone-based fluorescence imaging to yield an inexpensive yet ultrasensitive point-of-use diagnostic tool. Bioseparations dominate the cost and process complexity of manufacturing of modern biopharmaceuticals; process analytical (PAT) technologies that do not require trained investigators will immediately impact this area. In addition, a field-portable analytical method may prove useful for environmental contamination, food safety monitoring, and medical diagnostics. The University of Houston is the second-most ethnically diverse research university in the US and is designated as a Hispanic-serving Institution, providing a fertile recruiting ground for broadening participation from underrepresented groups. The PIs will continue ongoing outreach efforts aimed at the local scientific community, by organizing the Texas Soft Matter Meeting for academic and industrial scientists, and at society at large, by writing and presenting popular science radio programs for The Academic Minute and Engines of our Ingenuity.