We report a highly sensitive optical imaging technology using surface-enhanced Raman scattering (SERS)-fluorescence dual modal nanoprobes (DMNPs). Fluorescence microscopy is a well-known imaging technique that shows specific protein distributions within cells. However, most currently available fluorescent organic dyes have relatively weak emission intensities and are rapidly photo-bleached. Thus more sensitive and stable probes are needed. In this work we develop DMNPs, which can be used for both SERS and fluorescence detection. SERS detection is a powerful technique that allows ultrasensitive chemical or biochemical analysis through unlimited multiplexing and single molecule sensitivity. Combining advantages of fluorescence and SERS allows these dual modal nanostructures to be used as powerful probes for novel biomedical imaging. In this work, the fabrication and characterization of the SERS-fluorescence DMNPs and application to biological imaging were investigated using markers CD24 and CD44, which are co-expressed in MDA-MB-231 breast cancer cells, as a model system. SERS imaging with DMNPs was found to be a powerful tool to determine the co-localization of CD24 and CD44 in the cell.

Angiogenin, a 14-kDa protein first isolated from HT-29 human colon adenocarcinoma cells (1), is a potent blood vessel inducer present in mammalian plasma and milk (2)(3)(4). It has a unique ribonucleolytic activity (5,6) that is essential for neovascularization. It also stimulates endothelial cells to form diacylglycerol (7) and to secrete prostacyclin (8) by activating phospholipase C and phospholipase A2, respectively. It binds to calf pulmonary artery endothelial (CPAE) cells (9) and has been found to modulate a mitogenic effect in certain other cells (10). When angiogenin is modified by mutagenesis (11) or by limited proteolysis (12) in the region of residues 61-67, it loses angiogenin activity and will not compete with the native protein in the chicken embryo chorioallantoic membrane (CAM) assay. In contrast, angiogenin modified at either of the active-site histidine residues involved in ribonucleolytic activity, although inactive itself on the CAM, will compete with the native protein in this angiogenesis assay (13). These second-messenger binding and modulatory effects have led to the postulation of a dual-site model for the organogenic activity of angiogenin (12), which assumes that angiogenin interacts directly with cells, presumably through a membrane receptor, to exert this biological function.We report here the identification, isolation, and initial characterization of a bovine endothelial cell angiogeninbinding protein (AngBP) whose properties are consistent with its being a component of a cellular receptor of angiogenin.MATERIALS AND METHODS Materials. Bovine angiogenin, isolated from bovine milk as described (4), migrated as a single band in SDS/PAGE, induced vascularization in the chicken embryo CAM assay, and stimulated a second-messenger response in endothelial cells. Single-site mutants H13A and H114A in which the ribonucleolytic active-site histidine residues are changed to alanine,t the regional mutant ARH-Ij native human angiogenin, placental ribonuclease inhibitor (PRI), and bovine angiogenin were provided by R. Shapiro. The two proteolytic derivatives of human angiogenin, angiogenin K (cleaved at residues 60-61) and angiogenin E (cleaved at residues 67-68) were from T. Hallahan (12 2227The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

We report the rapid and highly sensitive trace analysis of mercury(ii) ions in water using a surface-enhanced Raman scattering (SERS)-based microdroplet sensor. Aptamer-modified Au/Ag core-shell nanoparticles have been fabricated and utilized as highly functional sensing probes. All detection processes for the reaction between mercury(II) ions and aptamer-modified nanoparticles were performed in a specially designed microdroplet channel. Small water droplets that included sample reagents were separated from each other by an oil phase that continuously flowed along the channel. This two-phase liquid-liquid segmented flow system prevented the adsorption of aggregated colloids to the channel walls due to localized reagents within encapsulated droplets. The result was reduced residence time distributions. The limit of detection (LOD) of mercury(II) ions in water was determined by the SERS-based microdroplet sensor to be below 10 pM, which is three orders below the EPA-defined maximum contaminant level. This combination of a SERS-based microfluidic sensor with aptamer-based functional nanoprobes can be used for in-the-field sensing platforms, due to its size and simplicity.

A surface-enhanced Raman scattering (SERS)-based gradient optofluidic sensor has been developed for a fast and sensitive immunoassay. In this work, a novel microfluidic sensor with functional internal structures has been designed and fabricated. This sensor is composed of three compartments consisting of the gradient channel that serially dilutes the target marker, the injection and mixing area of antibody-conjugated hollow gold nanospheres and magnetic beads, and the trapping area of sandwich immunocomplexes using multiple solenoids. Quantitative analysis of a specific target marker is performed by analyzing its characteristic SERS signals. This SERS-based gradient optofluidic sensor can replace the set of microwells or microtubes used in manual serial dilutions that have been traditionally used in enzyme-linked immunosorbent assay (ELISA)-type assays. The limit of detection for rabbit immunoglobin (IgG) is estimated to be 1-10 ng/mL. This novel SERS-based optofluidic immunoassay system is expected to be a powerful clinical tool for the fast and sensitive medical diagnosis of a disease.