This article provides an overview of the development and application of plasmonic nanoprobes developed in our laboratory for biosensing and bioimaging. We describe the use of plasmonics surface-enhanced Raman scattering (SERS) gene probes for the detection of diseases using DNA hybridization to target biospecies (HIV gene, breast cancer genes etc.). For molecular imaging, we describe a hyperspectral surface-enhanced Raman imaging (HSERI) system that combines imaging capabilities with SERS detection to identify cellular components using Raman dye-labeled silver nanoparticles in cellular systems The detection of specific target DNA sequences associated with breast cancer using “molecular sentinel” nanoprobes and the use of a plasmonic nanosensor to monitor pH in single cells are presented and discussed. Plasmonic nanosensors and nanoprobes have been developed as sensitive and selective tolls for environmental monitoring, cellular biosensing, medical diagnostics and high throughput screnning.
In this paper, we describe the development and application of a pH-sensitive plasmonics-active fiber-optic nanoprobe suitable for intracellular bioanalysis in single living human cells using surface-enhanced Raman scattering (SERS) detection. The effectiveness and usefulness of SERS-based fiber-optic nanoprobes are illustrated by measurements of intracellular pH in HMEC-15/hTERT immortalized “normal” human mammary epithelial cells and PC-3 human prostate cancer cells. The results indicate that fiber-optic nanoprobe insertion and interrogation provide a sensitive and selective means to monitor cellular microenvironments at the single cell level.
We report development of a nanoparticle-based, X-ray-activated anticancer "nanodrug" composed of yttrium oxide (Y(2)O(3)) nanoscintillators, a fragment of the HIV-1 TAT peptide, and psoralen. In this formulation, X-ray radiation is absorbed by the Y(2)O(3) nanoscintillators, which then emit UVA light. Absorption of UVA photons by nanoparticle-tethered psoralen has the potential to cross-link adenine and thymine residues in DNA. UVA-induced cross-linking by free psoralen upon activation with UVA light has previously been shown to cause apoptosis in vitro and an immunogenic response in vivo. Studies using the PC-3 human prostate cancer cell line demonstrate that X-ray excitation of these psoralen-functionalized Y(2)O(3) nanoscintillators yields concentration-dependent reductions in cell number when compared to control cultures containing psoralen-free Y(2)O(3) nanoscintillators.
We describe the development and application of a co-functionalized nanoprobe and biodelivery platform combining a nuclear targeting peptide (NTP) for improved cellular uptake and intracellular targeting with p-mercaptobenzoic acid (pMBA) as a surface-enhanced Raman scattering (SERS) reporter for tracking and imaging. The nuclear targeting peptide, an HIV-1 protein-derived TAT sequence, has been previously shown to aid entry of cargo through the cell membrane via normal cellular processes, and furthermore, to localize small cargo to the nucleus of the cell. Previous work in our lab has verified cell uptake and distribution of the nanoprobes in clinically relevant mouse and human cell lines. In this work, two-dimensional SERS mapping was used to track the spatial and temporal progress of nanoparticle uptake in PC-3 human prostate cells and to characterize localization at various time points, demonstrating the potential for an intracellularly targeted multiplexed nanobiosensing system with excellent sensitivity and specificity. Silver nanoparticles co-functionalized with the TAT peptide showed greatly enhanced cellular uptake over the control nanoparticles lacking the targeting moiety. The ability to detect and monitor nanoprobe trafficking using SERS spectroscopy offers an improved alternative over previous tracking and detection methods such as light microscopy and fluorescence methods. The development of multifunctional nanoconstructs for intracellular delivery has potential clinical applications in early detection and selective treatment of disease in affected cells. Other applications include use in basic research aimed at understanding the inner workings of living cells and how they respond to chemical and biological stimuli.
A critical aspect for use of nanoprobes in biomedical research and clinical applications involves fundamental spatial and temporal characterization of their uptake and distribution in cells. Raman spectroscopy and two-dimensional Raman imaging were used to identify and locate nanoprobes in single cells using surface-enhanced Raman scattering detection. To study the efficiency of cellular uptake, silver nanoparticles functionalized with three different positive-, negative-, and neutrally charged Raman labels were co-incubated with cell cultures and internalized via normal cellular processes. The surface charge on the nanoparticles was observed to modulate uptake efficiency, demonstrating a dual function of the surface modifications as tracking labels and as modulators of cell uptake. These results indicate that the functionalized nanoparticle construct has potential for sensing and delivery in single living cells and that use of surface-enhanced Raman scattering for tracking and detection is a practical and advantageous alternative to traditional fluorescence methods.
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