Gold nanoparticles targeting epidermal growth factor receptor via antibody conjugation undergo molecular specific aggregation when they bind to receptors on cell surfaces, leading to a red shift in their plasmon resonance frequency. Capitalizing on this effect, we demonstrate the efficacy of the molecular specific photoacoustic imaging technique using subcutaneous tumor-mimicking gelatin implants in ex-vivo mouse tissue. The results of our study suggest that highly selective and sensitive detection of cancer cells is possible using multiwavelength photoacoustic imaging and molecular specific gold nanoparticles.The developments in the fields of nanotechnology and molecular biology provide a promising platform for detection of cancer at an asymptomatic stage. Bioconjugated nano contrast agents together with imaging techniques can satisfy the compelling need to reliably detect, diagnose and characterize cancer at an early stage. [1][2][3][4][5][6][7] Recently, gold nanoparticles (Au NPs) have gained popularity as nano-sized contrast agents 2,6,[8][9][10][11][12][13][14] for their well-developed bioconjugation protocols, 11,[15][16][17] biocompatibility 18,19 and ease of tuning the optical properties. [20][21][22] Immunotargeted gold nanoparticles have been used to enhance contrast in optical imaging techniques. 6,9,13,14 However, the penetration depth achievable with high resolution optical imaging techniques is limited to a few millimeters. Optical techniques utilizing incoherent light extend the penetration depth to several centimeters while spatial resolution is severely sacrificed. Therefore, an in vivo imaging technique that is sensitive in detecting Au NPs and capable of imaging deep lying structures is desired. Photoacoustic imaging [23][24][25] is a technique that can provide penetration depth on the order of centimeters if near-infrared (NIR) laser light is used. In the photoacoustic phenomenon, 26 electromagnetic energy in the form of light is absorbed and subsequently an acoustic wave is emitted. Using a wideband ultrasound detector the acoustic waves can be detected and spatially resolved to provide an image of the optical absorption properties of the internal tissue structure. [23][24][25] Gold nanoparticles have been used as contrast agents in photoacoustic imaging because of their unique optical absorption properties. 8,10,[27][28][29][30][31] Using three-dimensional (3D) tissue models, we previously demonstrated that highly selective detection of cancer could be achieved using molecular targeted gold nanoparticles and combined photoacoustic and ultrasound imaging. 8,32 In particular, the contrast in the photoacoustic images was attributed to the epidermal growth factor receptor (EGFR) 33,34 leading to plasmon resonance coupling between adjacent gold particles and a red-shift in their absorbance spectra 6,8,9,14 while the nontargeted or isolated gold nanoparticles have absorbance peak at around 520 nm. 8,35,36 In this paper, we demonstrate the efficacy of multiwavelength photoacoustic imagin...

Polyethylene glycol (PEG) surface coatings are widely used to render stealth properties to nanoparticles in biological applications. There is abundant literature on benefits of PEG coatings and their ability to reduce protein adsorption, to diminish non-specific interactions with cells, and to improve pharmacokinetics, but very little discussion of the limitations of PEG coatings. Here, we show that physiological concentrations of cysteine and cystine can displace methoxy-PEG-thiol molecules from the gold nanoparticle (GNP) surface that leads to protein adsorption and cell uptake in macrophages within 24 hours. Furthermore, we address this problem by incorporating an alkyl linker between the PEG and the thiol moieties that provides a hydrophobic shield layer between the gold surface and the hydrophilic outer PEG layer. The mPEG-alkyl-thiol coating greatly reduces protein adsorption on GNPs and their macrophage uptake. This has important implications for the design of GNP for biological systems.

Based on their size and unique electrical properties, carbon nanotubes offer the exciting possibility of developing ultrasensitive, electrochemical biosensors. In this study, we describe the construction of amperometric biosensors based on the incorporation of single-walled carbon nanotubes modified with enzyme into redox polymer hydrogels. The composite films were constructed by first incubating an enzyme in a single-walled carbon nanotube (SWNTs) solution and then cross-linking within a poly[(vinylpyridine)Os(bipyridyl)(2)Cl(2+/3+)] polymer film. Incorporation of SWNTs, modified with glucose oxidase, into the redox polymer films resulted in a 2-10-fold increase in the oxidation and reduction peak currents during cyclic voltammetry, while the glucose electrooxidation current was increased 3-fold to approximately 1 mA/cm(2) for glucose sensors. Similar effects were also observed when SWNTs were modified with horseradish peroxidase prior to incorporation into redox hydrogels.

In this study, we describe the construction of glucose biosensors based on an electrostatic layer-by-layer (LBL) technique. Gold electrodes were initially functionalized with negatively charged 11-mercaptoundecanoic acid followed by alternate immersion in solutions of a positively charged redox polymer, poly[(vinylpyridine)Os(bipyridyl)2Cl(2+/3+)], and a negatively charged enzyme, glucose oxidase (GOX), or a GOX solution containing single-walled carbon nanotubes (SWNTs). The LBL assembly of the multilayer films were characterized by UV-vis spectroscopy, ellipsometry, and cyclic voltammetry, while characterization of the single-walled nanotubes was performed with transmission electron microscopy, Raman spectroscopy, thermogravimetric analysis, and X-ray photoelectron spectroscopy. When the GOX solution contained single-walled carbon nanotubes (GOX-SWNTs), the oxidation peak currents during cyclic voltammetry increased 1.4-4.0 times, as compared to films without SWNTs. Similarly the glucose electro-oxidation current also increased (6-17 times) when SWNTs were present. By varying the number of multilayers, the sensitivity of the sensors could be controlled.

Anisotropic gold nanorods provide a convenient combination of properties, such as tunability of plasmon resonances and strong extinction cross-sections in the near-infrared to red spectral region. These properties have created significant interest in the development of antibody conjugation methods for synthesis of targeted nanorods for a number of biomedical applications, including molecular specific imaging and therapy. Previously published conjugation approaches have achieved molecular specificity. However, the current conjugation methods have several downsides including low stability and potential cytotoxicity of bioconjugates that are produced by electrostatic interactions as well as lack of control over antibody orientation during covalent conjugation. Here we addressed these shortcomings by introducing directional antibody conjugation to the gold nanorod surface. The directional conjugation is achieved through the carbohydrate moiety, which is located on one of the heavy chains of the Fc portion of most antibodies. The carbohydrate is oxidized under mild conditions to a hydrazide reactive aldehyde group. Then, a heterofunctional linker with hydrazide and dithiol groups is used to attach antibodies to gold nanorods. The directional conjugation approach was characterized using electron microscopy, zeta potential and extinction spectra. We also determined spectral changes associated with nanorod aggregation; these spectral changes can be used as a convenient quality control of nanorod bioconjugates. Molecular specificity of the synthesized antibody targeted nanorods was demonstrated using hyperspectral optical and photoacoustic imaging of cancer cell culture models. Additionally, we observed characteristic changes in optical spectra of molecular specific nanorods after their interactions with cancer cells; the observed spectral signatures can be explored for sensitive cancer detection.