Electron tomography simulator with realistic 3D phantom for evaluation of acquisition, alignment and reconstruction methods.

Because of the significance of electron microscope tomography in the investigation of biological structure at nanometer scales, ongoing improvement efforts have been continuous over recent years. This is particularly true in the case of software developments. Nevertheless, verification of improvements delivered by new algorithms and software remains difficult. Current analysis tools do not provide adaptable and consistent methods for quality assessment. This is particularly true with images of biological samples, due to image complexity, variability, low contrast and noise. We report an electron tomography (ET) simulator with accurate ray optics modeling of image formation that includes curvilinear trajectories through the sample, warping of the sample and noise. As a demonstration of the utility of our approach, we have concentrated on providing verification of the class of reconstruction methods applicable to wide field images of stained plastic-embedded samples. Accordingly, we have also constructed digital phantoms derived from serial block face scanning electron microscope images. These phantoms are also easily modified to include alignment features to test alignment algorithms. The combination of more realistic phantoms with more faithful simulations facilitates objective comparison of acquisition parameters, alignment and reconstruction algorithms and their range of applicability. With proper phantoms, this approach can also be modified to include more complex optical models, including distance-dependent blurring and phase contrast functions, such as may occur in cryotomography.

Nonlinear immunofluorescent assay for androgenic hormones based on resonant structures.

We report for the first time the use of two photon fluorescence as detection method of affinity binding reactions. We use a resonant grating waveguide structure as platform enhancement for detecting the interaction between fluorescent labeled Boldenone, a non-natural androgenic hormone, and a specific anti-anabolic antibody. We were able to detect a surface coverage of approximately 0.7 ng/mm(2).

Iron-specific Signal Separation from within Heavy Metal Stained Biological Samples Using X-Ray Microtomography with Polychromatic Source and Energy-Integrating Detectors.

Biological samples are frequently stained with heavy metals in preparation for examining the macro, micro and ultra-structure using X-ray microtomography and electron microscopy. A single X-ray microtomography scan reveals detailed 3D structure based on staining density, yet it lacks both material composition and functional information. Using a commercially available polychromatic X-ray source, energy integrating detectors and a two-scan configuration labelled by their energy- "High" and "Low", we demonstrate how a specific element, here shown with iron, can be detected from a mixture with other heavy metals. With proper selection of scan configuration, achieving strong overlap of source characteristic emission lines and iron K-edge absorption, iron absorption was enhanced enabling K-edge imaging. Specifically, iron images were obtained by scatter plot material analysis, after selecting specific regions within scatter plots generated from the "High" and "Low" scans. Using this method, we identified iron rich regions associated with an iron staining reaction that marks the nodes of Ranvier along nerve axons within mouse spinal roots, also stained with osmium metal commonly used for electron microscopy.

Light modulation with electro-optic polymer-based resonant grating waveguide structures.

A novel modulator design incorporating an E-O polymer into a resonant grating waveguide structure is presented. Using purely polymeric material we developed a resonant grating waveguide structure having low loss and high finesse, with approximately 2nm spectral line width at 1.55 mum. An externally applied voltage modulates the refractive index of the E-O waveguide, thereby shifting the resonance wavelength and modulating the incident light at MHz rates. Such modulator operates in free space and does not involve waveguide patterning nor the need for facet conditioning and coupling common to operation in the Mach-Zehnder type configuration.

Role of photonic bandgaps in polarization-independent grating waveguide structures.

Polarization independence in a one-dimensional resonant grating waveguide structure involves the simultaneous excitation of two guided modes propagating in different directions. Possible simultaneous excitations occur when the two excited guided modes have either the same polarization, i.e., TE-TE (transverse electric) or TM-TM (transverse magnetic), or different polarizations, i.e., TE-TM. Simultaneous excitations may result in bandgaps and singularities. We confirm and show that in order to achieve polarization independence, it is necessary to find the conditions that minimize the effects of such bandgaps and singularities and experimentally demonstrate tunable polarization independence for simultaneously excited TE-TM-guided modes.