MicroRNAs (miRNAs) have increasingly become an important biomarker target for applications ranging from clinical diagnostics to biofuel production monitoring. However, the current state of the art for the detection of such markers requires tedious processing and amplification techniques such as polymerase chain reaction (PCR). In an effort to create a relatively simple biosensing platform, we have developed a combined plasmonic biosensing method based on a Surface-Enhanced Raman Spectroscopy (SERS) platform called the inverse Molecular Sentinel (iMS) to directly detect in vivo miRNA such as miR858a. With Shifted Excitation Raman Difference Spectroscopy (SERDS), we can remotely detect these targets in the field in the presence of interfering background signal. The application of such technology can pave the way not just for biofuel monitoring but early and non-invasive disease detection and diagnostics.
Plant biotechnology and biofuel research is critical in addressing increasing global demands for energy. Further understanding of biomass producing associated metabolic pathways in plants can be used to exploit and increase the production of biomass for energy purposes. In vivo detection of biomarkers associated with plant growth for bioenergy has proved to be limited due to complex sample preparation required by traditional methods. In addition, genetic transformation and biomolecule monitoring inside plant cells is regulated by diameter and size exclusion limits of the plant cell wall (5 - 20 nm). Currently limited methods exist for enabling direct entry into plant cells. Moreover, these methods, such as biolistic particle delivery and electroporation use mechanical force that causes damages to the plant tissue. Nanoparticles could serve as promising platforms for probes to characterize intercellular and intracellular plant biomarkers and pathways. Bi-metallic nanostars are a plasmonics-active nanoplatform capable of high surface-enhanced Raman scattering (SERS) which can enter plant cells and have the future potential for nucleic acid sensing. Imaging technologies such as SERS mapping, confocal imaging, X-ray fluorescence imaging, multi-photon imaging, and transmission electron microscopy have been utilized to determine the compartmentalization and location of the SERS iMS biosensors inside Arabidopsis plants.
Surface-enhanced Raman scattering (SERS) is emerging as an alternative non-invasive detection method in many applications. We recently show the use of SERS nanoprobes to detect tumors in vivo in mice, as well as the use of SERS sensors in vivo in plants for the detection of mIRNA. In spite of these advancements, the translation of SERS to real-world settings has been limited due to issues with observing Raman signal over complex background. For example, it remains challenging to observe SERS under sunlight or under strong illumination (e.g., operating room), using a conventional Raman setup. To this end, we combined a Raman setup with a newly developed dual-wavelength laser to perform shifted-excitation Raman difference spectroscopy (SERDS). Using SERDS, we demonstrate that the use of SERS sensors to detect miRNA in live plants inside a growth chamber, under full illumination. Additionally, we show that SERDS can be used to accurately identify tumors in mice, under ambient light. In both these applications, we demonstrate that the combination of SERS with SERDS improves the sensitivity and accuracy. This work will aid the translation of Raman and SERS to real-world settings.
The discovery of new treatments for cancer is imperative. Recently, we showed in a proof-of-concept study that SYnergistic IMmuno PHOtothermal NanotherapY (SYMPHONY) is a powerful treatment for metastatic bladder cancer and brain tumor in mouse models. Our work has recently demonstrated that combining immunotherapy checkpoint inhibitors and gold nanostar (AuNS) photothermal therapy (PTT) is more effective in killing primary tumors and activating the immune system to eradicate tumors at distant sites, than each individual treatment alone. When the tumor is being ablated via PTT in mice models, using low intensity heat from a near infrared laser, the dying tumor releases proteins that trigger the immune system to destroy remaining tumor cells. Immune checkpoint inhibitors prevent the tumor cells from hiding from the immune system’s mechanisms; thus, the immune system becomes capable of attacking distant secondary tumors, after the primary tumor has been eradicated using AuNS mediated PTT. The data shows that after the cured mice were rechallenged with bladder cancer cells, no tumor formation was observed after 60 days; hence these results indicate that the SYMPHONY treatment can function as a cancer vaccine and lead to long-lasting immunity.
Further understanding of biomass producing associated metabolic pathways in plants can be used to increase the production of biomass. In vivo detection of these markers has proved to be limited due to complex sample preparation required by traditional methods. Recently the Vo-Dinh group has designed a platform to detect nucleic acid targets in biological systems called inverse molecular sentinels which utilize surface-enhanced Raman scattering. These multimodal probes were shown to detect and image key microRNA within whole plants in vivo. This work lays the foundation for detecting and imaging biological markers in plants with enhanced spatial and temporal resolution.
Our group has integrated surface-enhanced Raman scattering (SERS) silver coated gold nanostars on an optical fiber. Fiber-based sensors are an in-situ technology that can simultaneously bring the sensor and light to the sample without disturbing the environment. This technology is a multi-use method that does not require complex sample preparation. Fiber sensors or optrodes, enable the detection of analytes in samples that are difficult to access. Additionally, optrodes allow for specific detection while evading background signals from non-target regions. The fiber-optrode was used to detect miRNA and illegal food additives.
Molecular analysis has revolutionized many applications, including bio-safety, bio-engineering and biofuel research; however, there are limited practical tools for in situ detection during field work. New technology is needed to translate molecular advances from laboratory settings into the practical realm. The unique characteristics of plasmonic nanosensors have made them ideal candidates for field-ready sensing applications. Herein, we discuss the development of a fiber-based plasmonic sensor capable of direct detection (i.e., no washing steps required) of miRNA targets, which are detected by immerging the sensor in the sample solution. This sensor is composed of an optical fiber that is decorated with plasmonic nanoprobes based on silver-coated gold nanostars to detect target nucleic acids using the surface-enhanced Raman scattering sensing mechanism of nanoprobes referred to as inverse molecular sentinels. The fiber sensors were tested in extracts from leaves of plants that were induced to have different miRNA expression levels. The results indicate that the fiber sensors developed have the potential to be a powerful tool for field analysis.
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