Fluorescence for biological logic gates.

Biological logic gates are smart probes able to respond to biological conditions in behaviors similar to computer logic gates, and they pose a promising challenge for modern medicine. Researchers are creating many kinds of smart nanostructures that can respond to various biological parameters such as pH, ion presence, and enzyme activity. Each of these conditions alone might be interesting in a biological sense, but their interactions are what define specific disease conditions. Researchers over the past few decades have developed a plethora of stimuli-responsive nanodevices, from activatable fluorescent probes to DNA origami nanomachines, many explicitly defining logic operations. Whereas many smart configurations have been explored, in this review we focus on logic operations actuated through fluorescent signals. We discuss the applicability of fluorescence as a means of logic gate implementation, and consider the use of both fluorescence intensity as well as fluorescence lifetime.

Gold Rod-Polyethylene Glycol-Carbon Dot Nanohybrids as Phototheranostic Probes.

Emphasis using phototheranostics has been placed on the construction of multifunctional nanoplatforms for simultaneous tumor diagnosis and therapy. Herein, we put forth a novel nanosized luminescent material using the incorporation of red emissive carbon dots on gold nanorods through polyethylene glycol as a covalent linkage for dual-modal imaging and photothermal therapy. The novel nanohybrids, not only retain the optical properties of the gold nanorod and carbon dots, but also possess superior imaging performance in both confocal laser scanning microscopy and fluorescence lifetime imaging microscopy. The nanohybrids also exhibit excellent photothermal performance as phototheranostic nanohybrid probes for in vitro assays. This study promises a new multifunctional nanoplatform for cancer diagnostics and therapeutics.

Development of a molecular bioswitch using fluorescence lifetime imaging: Incremental activation of fluorescein diacetate.

Molecular bioswitches offer an invaluable asset in the shift from systemic to targeted treatments. Within the growing arsenal of switches are imaging probes that functionalize only in given locations or situations. Acetate esters are a common fluorescent example, known to activate upon interaction with esterases. Fluorescein diacetate (FDA) is one such fluorophore used in cell viability assays. These assays rely on the fact that the compound begins colorless and with no fluorescent signature whatsoever, and only after internalization into cells it is possible to detect a fluorescence signal. In this study, using fluorescence intensity (FI) and fluorescence lifetime (FLT) imaging, FDA is shown to be fluorescent even when unactivated. Furthermore, the FLT is shown to change with pH. Finally, the ability to image FDA in different environments simulated by tissue-imitating phantoms is explored. Altogether, the ability of FDA to serve as a bioswitch when measured using FLT imaging microscopy (FLIM) is assessed. The combination of a spectrum of FDA activation and FLIM serves as a bioswitch, where biologically relevant stimulation can generate detectable and incremental variations.

Extremely sensitive dual imaging system in solid phantoms.

Herein we describe promising results from the combination of fluorescent lifetime imaging microscopy (FLIM) and diffusion reflection (DR) medical imaging techniques. Three different geometries of gold nanoparticles (GNPs) were prepared: spheres of 20nm diameter, rods (GNRs) of aspect ratio (AR) 2.5, and GNRs of AR 3.3. Each GNP geometry was then conjugated using PEG linkers estimated to be 10nm in length to each of 3 different fluorescent dyes: Fluorescein, Rhodamine B, and Sulforhodamine B. DR provided deep-volume measurements (up to 1cm) from within solid, tissue-imitating phantoms, indicating GNR presence corresponding to the light used by recording light scattered from the GNPs with increasing distance to a photodetector. FLIM imaged solutions as well as phantom surfaces, recording both the fluorescence lifetimes as well as the fluorescence intensities. Fluorescence quenching was observed for Fluorescein, while metal-enhanced fluorescence (MEF) was observed in Rhodamine B and Sulforhodamine B - the dyes with an absorption peak at a slightly longer wavelength than the GNP plasmon resonance peak. Our system is highly sensitive due to the increased intensity provided by MEF, and also because of the inherent sensitivity of both FLIM and DR. Together, these two modalities and MEF can provide a lot of meaningful information for molecular and functional imaging of biological samples.

Tissue-Like Phantoms as a Platform for Inserted Fluorescence Nano-Probes.

Tissue-like phantoms are widely used as a model for mimicking the optical properties of live tissue. This paper presents the results of a diffusion reflection method and fluorescence lifetime imaging microscopy measurements of fluorescein-conjugated gold nanorods in solution, as well as inserted in solid tissue-imitating phantoms. A lack of consistency between the fluorescence lifetime results of the solutions and the phantoms raises a question about the ability of tissue-like phantoms to maintain the optical properties of inserted contrast agents.

Targeted Magnetic Nanoparticles for Mechanical Lysis of Tumor Cells by Low-Amplitude Alternating Magnetic Field.

Currently available cancer therapies can cause damage to healthy tissue. We developed a unique method for specific mechanical lysis of cancer cells using superparamagnetic iron oxide nanoparticle rotation under a weak alternating magnetic field. Iron oxide core nanoparticles were coated with cetuximab, an anti-epidermal growth factor receptor antibody, for specific tumor targeting. Nude mice bearing a head and neck tumor were treated with cetuximab-coated magnetic nanoparticles (MNPs) and then received a 30 min treatment with a weak external alternating magnetic field (4 Hz) applied on alternating days (total of seven treatments, over 14 days). This treatment, compared to a pure antibody, exhibited a superior cell death effect over time. Furthermore, necrosis in the tumor site was detected by magnetic resonance (MR) images. Thermal camera images of head and neck squamous cell carcinoma cultures demonstrated that cell death occurred purely by a mechanical mechanism.

Validating a new computed tomography atlas for grading ankle osteoarthritis.

As the most common joint disease, osteoarthritis (OA) poses a significant source of pain and disability. It can be defined by classic radiographic findings, particular symptoms, or a combination of the 2. Although specific grading scales have been developed to evaluate OA in various joints, such as the shoulder, hip, and knee, no definitive classification system is available for grading OA in the ankle. The purpose of the present study was to create and validate a standardized atlas for grading (or staging) ankle osteoarthritis using computed tomography (CT) and "hallmark" findings noted on coronal, sagittal, and axial views extrapolated from the Kellgren-Lawrence radiographic scale. The CT scans of 226 patients at the Miami Veterans Affairs Medical Center were reviewed. An atlas was derived from a retrospective review of 30 remaining CT scans taken from July 2008 to November 2011. After this review, 3 orthogonal static CT images, obtained from 11 remaining patients, were chosen to represent the various stages on the OA scale and were used to test the validity of the atlas developed by 2 of us (M.M.C. and N.D.V.). A multispecialty panel of 9 examiners, excluding ourselves, independently rated the 11 CT scan subjects. The differences among examiners and specialties were calculated, including an intra-examiner agreement for 2 separate readings spaced 9 months apart. Although the small number of subspecialty examiners made the intraspecialty comparisons difficult to validate, the findings nevertheless indicated excellent agreement among all specialty groups, with good intra-investigational (intraclass correlation coefficient 0.962 and 1) inter-investigational (intraclass correlation coefficient 0.851) values. These results appeared to validate the CT ankle OA atlas, which we believe will be a valuable clinical and research tool, one that will likely be more beneficial than less relevant generalized OA grading scales in use today.

An ultra-sensitive dual-mode imaging system using metal-enhanced fluorescence in solid phantoms.

In this study we developed a highly sensitive dual modal imaging system designed for gold nanoparticles (GNPs) conjugated to various fluorophores in solid phantoms. The system consists of fluorescence lifetime imaging microscopy (FLIM) for surface imaging, diffusion reflection (DR) for deep tissue imaging (up to 1cm), and metal enhanced fluorescence (MEF). We detected quenching in fluorescent intensity (FI) for the conjugation of gold nanospheres (GNS) as well as gold nanorods (GNRs) to Fluorescein, which has an excitation peak at a wavelength shorter than the surface plasmon resonance (SPR) of both types of GNPs, and enhanced FI in conjugation to Rhodamine B and Sulforhodamine B, both with excitation peaks in the GNPs' SPR. The enhanced FI was detected in solution as well as in solid phantoms from FLIM measurements. DR measurements detected GNR presence within the solid phantoms by recording dropped rates of light scattering using wavelengths corresponding to the GNRs' absorption. With the inclusion of MEF, this promising dual modal imaging technique enables efficient and sensitive molecular and functional imaging.