Browsing by Subject "Plant Biotechnology"

In this thesis, we present an overview of the development and application of surface-enhanced Raman scattering (SERS) and plasmonic nanoplatforms developed in our laboratory for sensing applications. Plasmonic nanomaterials, such as gold nanostars (AuNS) a hallmark particle pioneered by the Vo-Dinh group, increase inherently weak Raman signals from molecules providing an intense and unique SERS spectrum allowing for targets to be sensitively detected and easily identified. For this work, we investigated biosensing in plants and murine models using plasmonic nanoplatforms. In the first part of the work, we developed a wide variety of plasmonics-active substrates and nanoparticle-based sensing systems including inverse molecular sentinels (iMS) utilizing two platforms silver-coated gold nanostars (AuNS@Ag) and nanorods (AuNR@Ag). The AuNS@Ag were decorated onto fiber-optrodes for chemical fiber sensing and monitoring of genomic biomarkers in plants for renewable bioenergy research. Rapid chemical sensing of illegal food additives proves to be a challenge at the site of exposure thus requiring the need for in situ fiber detection. Also, the fiber sensing approach is necessary for facilitating field analyses of microRNAs since the gold standard methods can only be performed in laboratory settings on the timescale of days. The fiber-optrodes were capable of functioning as chemical and biological sensors for analytes such as illegal food additives and plant microRNAs. The work is also aimed at microRNA sensing but directly inside of plant tissue with spatial and temporal resolutions that PCR cannot achieve. Thus, the silver-coated nanorod (AuNR@Ag) was developed with the purpose of infiltrating plant cells. We designed the plasmonic nanorod to have a dimension smaller than the plant cell wall exclusion limit to permit cellular uptake, while improving SERS properties through a silver coating on the particle. We confirmed particle uptake in plant cells using a multi-modal approach consisting of confocal microscopy, transmission electron microscopy, and x-ray fluorescence microscopy. Dye coated AuNR@Ag served as a strong contrast agent for two-photon imaging, photoacoustic imaging, and Raman mapping during in vivo experiments in Tobacco leaves. The AuNR@Ag was further functionalized with iMS technology and was applied for sensing for microRNA targets in leaf tissue. To our knowledge, this is the first demonstration of intracellular SERS sensing in vivo of leaf tissue treated with the AuNR@Ag nanoprobes. In another chapter of the work, we used standoff shifted-excitation difference spectroscopy (SERDS) for remote detection of biomarkers in plants under ambient light conditions. Monitoring plant molecular targets in field conditions remain an elusive task for standard optical methods such as fluorescence and Raman spectroscopy. However, recent developments in nanoprobe technology and remote optical techniques have ushered in a novel mechanism for highly specific molecular monitoring of living organisms at different stages of growth and other phenotypic cues. We have successfully demonstrated nanoprobe detection in a live plant leaf at a minimal distance of 2 meters. This work brings the remote monitoring of plant genetic biomarkers closer to in vivo tracking and analysis without the need of a dark laboratory as required by traditional optical sensing. Applications of plasmonic AuNS for bioimaging of tumors in combination with SERDS is also presented. During surgery accurately removing the entire tumor without harming surrounding healthy tissue is critical; however, due to the lack of intraoperative imaging techniques, surgeons rely on visual and physical inspection to identify tumors. To address this problem, we established the first use of SERDS for in vivo tumor detection in a murine model under ambient light conditions, mimicking an intraoperative environment.