Nanoplasmonic spectroscopic imaging and molecular probes for living cells

Label-free, sensitive and selective detection methods with high spatial resolution are critically required for future applications in chemical sensor, biological sensor, and nanospectroscopic imaging. Fluorophore or quantum dot-based detection are facing their limit due to photobleaching and blinking, and plasmonic-based sensing methods such as Surface Plasmon Resonance (SPR), Localized Surface Plasmon Resonance (LSPR), and Surface Enhanced Raman Spectroscopy (SERS) are limited in spatial resolution, molecular finger print information, or uniformity of detection. However, these limits can be overcome by using the recently introduced plasmon based detection method, Plasmon Resonance Energy Transfer (PRET).Here I describe the development of PRET-based molecular imaging in living cells as the first demonstration of intracellular imaging with PRET-based nanospectroscopy. In-vivo PRET imaging relied on the overlap between plasmon resonance frequency of gold nanoplasmonic probe (GNP) and absorption peak frequencies of conjugated molecules, which leads to create 'quantized quenching dips' in Rayleigh scattering spectrum of GNP. The position of these dips exactly matched with the absorption peaks of target molecules. Such the dips allow quantitative and long-term dynamic imaging with nanoscale spatial resolution.As another innovative application of PRET, I present a highly selective and sensitive detection of metal ions by creating conjugated metal-ligand complexes on a single GNP. In addition to conferring high spatial resolution due to the small size of the metal ion probes (50 nm in diameter), this method is 100 to 1,000 folds more sensitive than organic reporter-based methods. Moreover, this technique achieves high selectivity due to the selective formation of Cu2+ complexes and selective resonant quenching of GNP by the conjugated complexes. Since many metal ion ligand complexes generate new absorption peak due to the d-d transition in the metal ligand complex when a specific metal ion is inserted into the complex, we can match with the scattering frequency of nanoplasmonic metal ligand systems and the new absorption peak. The PRET-based metal ion detection method is easily expanded to detect other metal ions by replacing the conjugated ligand on the surface of GNPs.In addition to in-vivo molecular imaging application nanoplasmonics, I also have developed more reliable and sensitive molecular probe arrays for in-vitro applications. In order to accomplish an integrated nanoplasmonic label-free bioassay array system, we need to overcome the limitation of current nanoscale batch-fabrication method. Classical optical lithography is limited by diffraction of light for nanolithography. Electron beam lithography and focused ion beam show serious challenge due to the inherent serial processing. As an alternative nanofabrication to combine the advantages of parallel processing, tunable capability of geometries, cost-effective method, and high spatial resolution nanolithography technique, As an innovative solution to reduce the current nanolithographic limit in terms of resolution, I have accomplished Shadow Angle Interference Lithography (SAIL). The SAIL relies on the overlap of shadows (i.e. shadow interference) created by the directional metal deposition angle and etching angle on pre-patterned template. As a result, highly tunable nanostructure array can be obtained.As an example of ordered nanoplasmonic array in large-area, I demonstrate the spontaneous formation of plasmonic nanocrown consisting of one spherical particle with 50 nm diameter at center (i.e. planet nanoparticle) and six smaller satellite particles around it. Well-spaced, high-density nanocrown is prepared via dewetting of gold thin film on anodic aluminum oxide (AAO) which has regular nano-pattern and it displays a frequency-selective response in the visible wavelengths depending on the distance (100 nm &sim 1mum) between the planet and satellite particles.The precisely and uniformly fabricated nanostructure array can be used as plasmonic based detection template and combine with microfluidic system for parallel and quantitative molecular and cellular experiments. This combined system can have a large impact on the fields of quantitative in-vivo cellular imaging, pharmacology, systems biology, and environmental monitoring.