Cooled Charge Coupled Device Camera Research Articles

Development of a novel delivery quality assurance system based on simultaneous verification of dose distribution and binary multi-leaf collimator opening in helical tomotherapy

BackgroundIntensity-modulated radiation therapy (IMRT) requires delivery quality assurance (DQA) to ensure treatment accuracy and safety. Irradiation techniques such as helical tomotherapy (HT) have become increasingly complex, rendering conventional verification methods insufficient. This study aims to develop a novel DQA system to simultaneously verify dose distribution and multi-leaf collimator (MLC) opening during HT.MethodsWe developed a prototype detector consisting of a cylindrical plastic scintillator (PS) and a cooled charge-coupled device (CCD) camera. Scintillation light was recorded using a CCD camera. A TomoHDA (Accuray Inc.) was used as the irradiation device. The characteristics of the developed system were evaluated based on the light intensity. The IMRT plan was irradiated onto the PS to record a moving image of the scintillation light. MLC opening and light distribution were obtained from the recorded images. To detect MLC opening, we placed a region of interest (ROI) on the image, corresponding to the leaf position, and analyzed the temporal change in the light intensity within each ROI. Corrections were made for light changes due to differences in the PS shape and irradiation position. The corrected light intensity was converted into the leaf opening time (LOT), and an MLC sinogram was constructed. The reconstructed MLC sinogram was compared with that calculated using the treatment planning system (TPS). Light distribution was obtained by integrating all frames obtained during IMRT irradiation. The light distribution was compared with the dose distribution calculated using the TPS.ResultsThe LOT and the light intensity followed a linear relationship. Owing to MLC movements, the sensitivity and specificity of the reconstructed sinogram exceeded 97%, with an LOT error of − 3.9 ± 7.8%. The light distribution pattern closely resembled that of the dose distribution. The average dose difference and the pass rate of gamma analysis with 3%/3 mm were 1.4 ± 0.2% and 99%, respectively.ConclusionWe developed a DQA system for simultaneous and accurate verification of both dose distribution and MLC opening during HT.

Feasibility study of optical imaging of the boron-dose distribution by a liquid scintillator in a clinical boron neutron capture therapy field.

Evaluation of the boron dose is essential for boron neutron capture therapy (BNCT). Nevertheless, a direct evaluation method for the boron-dose distribution has not yet been established in the clinical BNCT field. To date, even in quality assurance (QA) measurements, the boron dose has been indirectly evaluated from the thermal neutron flux measured using the activation method with gold foil or wire and an assumed boron concentration in the QA procedure. Recently, we successfully conducted optical imaging of the boron-dose distribution using a cooled charge-coupled device (CCD) camera and a boron-added liquid scintillator at the E-3 port facility of the Kyoto University Research Reactor (KUR), which supplies an almost pure thermal neutron beam with very low gamma-ray contamination. However, in a clinical accelerator-based BNCT facility, there is a concern that the boron-dose distribution may not be accurately extracted because the unwanted luminescence intensity, which is irrelevant to the boron dose is expected to increase owing to the contamination of fast neutrons and gamma rays. The purpose of this research was to study the validity of a newly proposed method using a boron-added liquid scintillator and a cooled CCD camera to directly observe the boron-dose distribution in a clinical accelerator-based BNCT field. A liquid scintillator phantom with 10 B was prepared by filling a small quartz glass container with a commercial liquid scintillator and boron-containing material (trimethyl borate); its natural boron concentration was 1wt%. Luminescence images of the boron-neutron capture reaction were obtained in a water tank at several different depths using a CCD camera. The contribution of background luminescence, mainly due to gamma rays, was removed by subtracting the luminescence images obtained using another sole liquid scintillator phantom (natural boron concentration of 0wt%) at each corresponding depth, and a depth profile of the boron dose with several discrete points was obtained. The obtained depth profile was compared with that of calculated boron dose, and those of thermal neutron flux which were experimentally measured or calculated using a Monte Carlo code. The depth profile evaluated from the subtracted images indicated reasonable agreement with the calculated boron-dose profile and thermal neutron flux profiles, except for the shallow region. This discrepancy is thought to be due to the contribution of light reflected from the tank wall. The simulation results also demonstrated that the thermal neutron flux would be severely perturbed by the 10 B-containing phantom if a relatively larger container was used to evaluate a wide range of boron-dose distributions in a single shot. This indicates a trade-off between the luminescence intensity of the 10 B-added phantom and its perturbation effect on the thermal neutron flux. Although a partial discrepancy was observed, the validity of the newly proposed boron-dose evaluation method using liquid-scintillator phantoms with and without 10 B was experimentally confirmed in the neutron field of an accelerator-based clinical BNCT facility. However, this study has some limitations, including the trade-off problem stated above. Therefore, further studies are required to address these limitations.

Technical note: Optical imaging of lithium‐containing zinc sulfate plate in water during irradiation of neutrons from boron neutron capture therapy (BNCT) system

Optical imaging of ionizing radiation is a possible method for dose distribution measurements. However, it is not clear whether the imaging method is also applicable to neutrons. To clarify this, we performed the imaging of neutrons in water from boron neutron capture therapy (BNCT) systems. Such systems require efficient distribution measurements of neutrons for quality assessment (QA) of the beams. A water-filled phantom was irradiated from the side with an epithermal neutron beam, in which a lithium-containing zinc sulfate (Li-ZnS(Ag)) plate was set in the beam direction, and during this irradiation the scintillation of the plate was imaged using a cooled charge-coupled device (CCD) camera. In the imaging, Li-6 in the Li-ZnS(Ag) plate captures neutrons and converts them to alpha particles (He-4) and tritium (H-3), while ZnS(Ag) in the Li-ZnS(Ag) plate produces scintillation light in the plate. We also conducted Monte Carlo simulation and compared its results with the experimental results. The image of the emitted light from the Li-ZnS(Ag) plate was clearly obtained with an imaging time of 0.5s. The depth and lateral profiles of the measured image using the Li-ZnS(Ag) plate showed the same shapes as the neutron distributions measured with gold foil, within a difference of 8%. The destructive effect of neutrons on the CCD camera increased approximately three times, but the unit was still working after the measurement. The optical imaging of neutrons in water is possible, and it has the potential to be a new method for efficient QA as well as for research on neutrons.

First optical observation of 10B-neutron capture reactions using a boron-added liquid scintillator for quality assurance in boron neutron capture therapy.

10B-neutron capture was observed optically using a boron-added liquid scintillator. Trimethyl borate was dissolved in a commercially available liquid scintillator at natural boron concentrations of approximately 1 wt% and 0.25 wt%. The boron-added liquid scintillator was placed in a phantom quartz bottle and irradiated by thermal neutrons (~ 105 n/[cm2 s]) for 150, 300, and 600s. The luminescence of the liquid scintillator was clearly observed using a cooled charge-coupled device (CCD) camera during irradiation. The luminance value recorded by the CCD camera was proportional to the duration of irradiation by thermal neutrons. The luminescence distribution showed reasonable agreement with that of energy deposition by Li and alpha particles from 10B-neutron capture reactions calculated via Monte Carlo simulations. When trimethyl borate was not dissolved in the liquid scintillator (0 wt% natural boron), no visible luminescence was observed even after 600s of irradiation. These findings demonstrate that the observed luminance originates from the Li and alpha particles generated by 10B-neutron capture reactions. Consequently, the luminescence distribution is directly related to the boron dose of the liquid scintillator. To the best of our knowledge, direct experimental optical observations of boron dose distribution have not yet been reported. This novel technique will be useful for quality assurance in boron neutron capture therapy (BNCT) because instantaneous neutron irradiation may be sufficient for the observing the intense neutron beam used in clinical BNCT (~ 109 n/[cm2 s]), and quick evaluation of the boron dose distribution is expected to be feasible.

A Smart Advanced Chemiluminescence-Sensing Platform for Determination and Imaging of the Tissue Distribution of Natural Antioxidants.

Antioxidants have gained marked attention owing to their ability to prevent the oxidation of biological components and to protect the body from reactive oxygen species, thereby maintaining human health. Thus, antioxidant-rich dietary supplements and natural foods can be effective against oxidative stress and can even act as chemopreventive agents. Therefore, a simple and rapid assay for evaluation of antioxidant capacity and assessment of their distribution profile in natural sources is vital. Herein, we report a rapid, innovative chemiluminescence (CL) platform for evaluation and visualization of antioxidant capacity. We found that intense and long-lasting CL was formed upon the redox reaction of quinones, e.g., menadione, with antioxidants, e.g., l-ascorbic acid, in the presence of luminol. The produced CL intensities were proportional to the antioxidants' concentrations with a detection limit of 0.18 μM for the model antioxidant, l-ascorbic acid. As the formed CL was long-lasting, it could be easily captured and detected with a charge-coupled device (CCD) camera. To evaluate the quantification ability of the CCD camera, we developed a smart and fast microplate-based assay based on photographing the generated CL with a cooled CCD camera. The photographed CL intensities were linearly proportional with the antioxidant concentrations, and then the method was applied for photographing multiple food sample extracts. Ultimately, we utilized our method for the distribution profiling of antioxidant capacity in food cut sections. Samples were dipped in luminol and then in quinone, followed by CCD camera photography, without the need for any pulverization/extraction procedure, giving precise antioxidant distribution information.

Imaging of fragment particles in water by nuclear spallation during carbon-ion irradiation

Recently we found that the luminescence imaging of water during carbon-ion irradiation was possible using a cooled charge-coupled device (CCD) camera and the method could be used for range estimation of the beam. In the luminescence image, we found luminescence from the fragment particles produced by the nuclear spallation reaction of carbon ions. The luminescence may be used for the estimation of the distribution of the fragment particles by the nuclear spallation. For this purpose, we irradiated carbon ions of 241.5 MeV u−1 to a water phantom and measured the luminescence image of water using a CCD camera. Then, we carefully observed the luminescence distribution after the Bragg peak to find the luminescence from the nuclear spallation reaction. In the luminescence image, we could clearly observe the luminescence from the fragment particles produced by the nuclear spallation reaction during irradiation of carbon ions. The beam widths of the luminescence image of the nuclear spallation were compared with those measured by the ionization chamber. The relative difference of the beam width at FWHM between luminescence image and ionization chamber was 23%. With these results, we conclude that the luminescence image of water during carbon-ion irradiation has a potential to be a new and efficient method for the width estimation of the fragment particles by the nuclear spallation reaction.

Luminescence imaging of water during uniform-field irradiation by spot scanning proton beams

Luminescence was found during pencil-beam proton irradiation to water phantom and range could be estimated from the luminescence images. However, it is not yet clear whether the luminescence imaging is applied to the uniform fields made of spot-scanning proton-beam irradiations. For this purpose, imaging was conducted for the uniform fields having spread out Bragg peak (SOBP) made by spot scanning proton beams. We designed six types of the uniform fields with different ranges, SOBP widths and irradiation fields. One of the designed fields was irradiated to water phantom and a cooled charge coupled device camera was used to measure the luminescence image during irradiations. We estimated the ranges, field widths, and luminescence intensities from the luminescence images and compared those with the dose distribution calculated by a treatment planning system. For all types of uniform fields, we could obtain clear images of the luminescence showing the SOBPs. The ranges and field widths evaluated from the luminescence were consistent with those of the dose distribution calculated by a treatment planning system within the differences of −4 mm and −11 mm, respectively. Luminescence intensities were almost proportional to the SOBP widths perpendicular to the beam direction. The luminescence imaging could be applied to uniform fields made of spot scanning proton beam irradiations. Ranges and widths of the uniform fields with SOBP could be estimated from the images. The luminescence imaging is promising for the range and field width estimations in proton therapy.

Chapter 9 - A High-Resolution Multimode Digital Microscope System

This chapter describes the development of a high-resolution, multimode digital imaging system based on a wide-field epifluorescent and transmitted light microscope, and a cooled charge-coupled device (CCD) camera. The three main parts of this imaging system are Nikon FXA microscope, Hamamatsu C4880 cooled CCD camera, and MetaMorph digital imaging system. This chapter presents various design criteria for the instrument and describes the major features of the microscope components-the cooled CCD camera and the MetaMorph digital imaging system. The Nikon FXA upright microscope can produce high resolution images for both epifluorescent and transmitted light illumination without switching the objective or moving the specimen. The functional aspects of the microscope set-up can be considered in terms of the imaging optics, the epi-illumination optics, the transillumination optics, the focus control, and the vibration isolation table. This instrument is somewhat specialized for microtubule and mitosis studies, and it is also applicable to a variety of problems in cellular imaging, including tracking proteins fused to the green fluorescent protein in live cells. The instrument is also valuable for correlating the assembly dynamics of individual cytoplasmic microtubules (labeled by conjugating X-rhodamine to tubulin) with the dynamics of membranes of the endoplasmic reticulum (labeled with DiOC6) and the dynamics of the cell cortex (by differential interference contrast) in migrating vertebrate epithelial cells. This imaging system also plays an important role in the analysis of mitotic mutants in the powerful yeast genetic system Saccharomyces cerevisiae.

Continuous sensing of tumor-targeted molecular probes with a vertical cavity surface emitting laser-based biosensor

Molecular optical imaging is a widespread technique for interrogating molecular events in living subjects. However, current approaches preclude long-term, continuous measurements in awake, mobile subjects, a strategy crucial in several medical conditions. Consequently, we designed a novel, lightweight miniature biosensor for in vivo continuous optical sensing. The biosensor contains an enclosed vertical-cavity surface-emitting semiconductor laser and an adjacent pair of near-infrared optically filtered detectors. We employed two sensors (dual sensing) to simultaneously interrogate normal and diseased tumor sites. Having established the sensors are precise with phantom and in vivo studies, we performed dual, continuous sensing in tumor (human glioblastoma cells) bearing mice using the targeted molecular probe cRGD-Cy5.5, which targets αVβ3 cell surface integrins in both tumor neovasculature and tumor. The sensors capture the dynamic time-activity curve of the targeted molecular probe. The average tumor to background ratio after signal calibration for cRGD-Cy5.5 injection is approximately 2.43±0.95 at 1 h and 3.64±1.38 at 2 h (N=5 mice), consistent with data obtained with a cooled charge coupled device camera. We conclude that our novel, portable, precise biosensor can be used to evaluate both kinetics and steady state levels of molecular probes in various disease applications.

Luciferase bioluminescence imaging monitoring gene therapeutic effect of apoptosis-inducing ligand for lung cancer A549 cells nude mice transplantation tumor in vivo

Objective To detect the expression and effect of human tumor necrosis factor related apoptosis-inducing ligand (hTRAIL) in vivo,by using a novel double expressing adenoviral vector encoding hTRAIL and firefly luciferase ( luc ) gene ( ad -luc-hTRAIL),in which luc was used as reporter gene.Methods Lung cancer A549 cell xenografts in 16 nude mice models were established in subcutaneous inoculation way,the adenovirus vectors ( ad-luc-hTRAIL,ad-hTRAIL,ad-luc) and phosphate buffer saline (PBS) (n =4) as control were injected into tumor respectively.The size of the tumor was measured at different time points (4,7,10,14,21,28 d)after injection.The activity of luciferase in surface of the tumor was detected in vivo by using high-sensitivity cooled-charged coupled device(CCD) camera.The expression of hTRAIL was demonstrated by immunohistochemistry staining after sacrificing the animals at different time points,and immunohistochemical scores (IHS) were measured. The apoptosis rate of tumor cells was detected by using TUNEL and calculated. Analysis of variance,the paired t test and linear correlation analysis was used for the statistics. Results The growing speed of tumour xenografts was more slowly in ad-luc-hTRAIL and ad-hTRAIL groups than PBS group (t =2.71,2.72,P ＜ 0.05 ).The tumor volumes of ad-luc-hTRAIL,ad-hTRAIL,ad-luc and PBS groups 28 days after injection were (208.4 ± 42.3 ),( 181.5 ±23.9),( 403.1 ± 54.0 ) and ( 427.0 ± 59.3 ) mm3, respectively. There was no significant difference between ad-luc group and PBS group(t =2.07,P ＞ 0.05).The expression of luciferase in ad-luc-hTRAILgroup reached its peak at 7th day ( 1.37 ± 1.04),and then decreased quickly.The IHS and apoptosis rate in ad-luc-hTRAIL and ad-hTRAIL groups reached their peaks at 7th day,the peak values of IHS were 6.25 ±2.06 and 6.5 ± 2.89,the peak values of apoptosis rate were (60.75 ± 8.06 ) ％ and ( 61.50 ± 8.47 ) ％,respectively.The amount of luciferase expression ( absolute number of photons detected by CCD camera)was linear positive correlated with IHS and apoptosis rate ( rphotons/IHs =0.942,rph /rate =0.842,rIHs/rate =0.887,P ＜ 0.05 ).Conclusion The target gene hTRAIL can be transfected into lung cancer A549 cell xenografts nude mice models efficiently with a high level expression,and the therapeutic effect of hTRAIL can be monitored by detecting the expression of luc. Key words: Luminescent proteins; Tumor necrosis factors; Fluorescein; Genes

A Water-Soluble Coelenterazine for Sensitive In Vivo Imaging of Coelenterate Luciferases

A Water-Soluble Coelenterazine for Sensitive In Vivo Imaging of Coelenterate Luciferases

Using spontaneous photon emission to image lipid oxidation patterns in plant tissues

Plants, like almost all living organisms, spontaneously emit photons of visible light. We used a highly sensitive, low-noise cooled charge coupled device camera to image spontaneous photon emission (autoluminescence) of plants. Oxidative stress and wounding induced a long-lasting enhancement of plant autoluminescence, the origin of which is investigated here. This long-lived phenomenon can be distinguished from the short-lived chlorophyll luminescence resulting from charge recombinations within the photosystems by pre-adapting the plant to darkness for about 2 h. Lipids in solvent were found to emit a persistent luminescence after oxidation in vitro, which exhibited the same time and temperature dependence as plant autoluminescence. Other biological molecules, such as DNA or proteins, either did not produce measurable light upon oxidation or they did produce a chemiluminescence that decayed rapidly, which excludes their significant contribution to the in vivo light emission signal. Selective manipulation of the lipid oxidation levels in Arabidopsis mutants affected in lipid hydroperoxide metabolism revealed a causal link between leaf autoluminescence and lipid oxidation. Addition of chlorophyll to oxidized lipids enhanced light emission. Both oxidized lipids and plants predominantly emit light at wavelengths higher than 600 nm; the emission spectrum of plant autoluminescence was shifted towards even higher wavelengths, a phenomenon ascribable to chlorophyll molecules acting as luminescence enhancers in vivo. Taken together, the presented results show that spontaneous photon emission imaged in plants mainly emanates from oxidized lipids. Imaging of this signal thus provides a simple and sensitive non-invasive method to selectively visualize and map patterns of lipid oxidation in plants.

A CMOS In-Pixel CTIA High-Sensitivity Fluorescence Imager

Traditionally, charge coupled device (CCD) based image sensors have held sway over the field of biomedical imaging. Complementary metal oxide semiconductor (CMOS) based imagers so far lack sensitivity leading to poor low-light imaging. Certain applications including our work on animal-mountable systems for imaging in awake and unrestrained rodents require the high sensitivity and image quality of CCDs and the low power consumption, flexibility and compactness of CMOS imagers. We present a 132×124 high sensitivity imager array with a 20.1 μm pixel pitch fabricated in a standard 0.5 μ CMOS process. The chip incorporates n-well/p-sub photodiodes, capacitive transimpedance amplifier (CTIA) based in-pixel amplification, pixel scanners and delta differencing circuits. The 5-transistor all-nMOS pixel interfaces with peripheral pMOS transistors for column-parallel CTIA. At 70 fps, the array has a minimum detectable signal of 4 nW/cm(2) at a wavelength of 450 nm while consuming 718 μA from a 3.3 V supply. Peak signal to noise ratio (SNR) was 44 dB at an incident intensity of 1 μW/cm(2). Implementing 4×4 binning allowed the frame rate to be increased to 675 fps. Alternately, sensitivity could be increased to detect about 0.8 nW/cm(2) while maintaining 70 fps. The chip was used to image single cell fluorescence at 28 fps with an average SNR of 32 dB. For comparison, a cooled CCD camera imaged the same cell at 20 fps with an average SNR of 33.2 dB under the same illumination while consuming over a watt.

Imaging Performance Improvement of an Extreme Ultraviolet Microscope

The extreme ultraviolet microscope (EUVM) has been developed for an actinic mask inspection of a EUV finished mask and a EUV blank mask. Using this microscope, amplitude defects on a finished mask and phase defects on a glass substrate are observed. However, it has a problem of low contrast, which originates from 1) thermal noise of a charge coupled device (CCD) camera, 2) wave aberrations of an optical component, and 3) a nonuniform illumination intensity. To resolve these issues, EUVM was improved. 1) To reduce a thermal noise, a cooled CCD camera is installed. 2) To remove wave aberrations of a back-end turning mirror, a Mo/Si multiplayer-coated thick glass substrate with a high surface accuracy is employed instead of a Si wafer substrate. Furthermore, in situ alignment was carried out to remove wavefront aberrations for a Schwarzschild imaging optics. In addition, 3) by installing a scanning system on the front-end turning mirror, a highly uniform illumination intensity was achieved. As a result, images of less than 100 nm without astigmatism were obtained.

Combined system of fluorescence diffuse optical tomography and microcomputed tomography for small animal imaging

We developed a dual-modality system that combines fluorescence diffuse optical tomography (fDOT) and flat panel detector-based microcomputed tomography (micro-CT) to simultaneously reveal molecular and structural information in small animals. In fDOT, a 748 nm diode laser was used as an excitation source, while a cooled charge coupled device camera was adopted to collect transmission fluorescence. In micro-CT, a flat panel detector based on amorphous silicon, with active area of 13 x 13 cm(2), and a microfocus x-ray tube were used. The fDOT system was mounted orthogonally to the micro-CT and the projection images were acquired without rotation of the sample, which is different from the method used for micro-CT alone. Both the finite element method and the algebraic reconstruction technique were used to reconstruct images from the fDOT. Phantom data showed that the resolution of the fDOT system was about 3 mm at an imaging depth of 7 mm. Quantitative error was no more than 5% and imaging sensitivity for 1,1(')-dioctadecyl-3,3,3('),3(')-etramethylindotricarbocyanine iodide bis-oleate (DiR-BOA) was estimated to be higher than 100 nM at a depth of 7 mm. Calculations of the phantom's center of mass showed that the location accuracy of fDOT was about 0.7 mm. We applied a Feldkamp algorithm to reconstruct the micro-CT image. By measuring the presampled modulation transfer function with a 30 microm tungsten thread, we estimated that the micro-CT has a resolution of 5 mm(-1) when the field of view was 6.5 cm. Our results indicate the uniformity of the transaxial micro-CT image and the contrast-to-noise ratio was measured as 1.95 for a radiation dose of 1 cGy. A non-image-based method was employed for merging images from the two imaging modalities. A nude mouse with DiR-BOA, imaged ex vivo, was used to validate the feasibility of the dual-modality system.

C-Jun N-terminal Kinase Is Necessary for Platelet-derived Growth Factor-mediated Chemotaxis in Primary Fibroblasts

c-Jun N-terminal kinase (JNK) is a member of the mitogen-activated protein kinase family. It has become clear that JNK does not only have a role in induction of stress responses but also in processes such as cell movement. In this report we demonstrate that JNK activity is necessary for platelet-derived growth factor (PDGF)-BB-induced chemotaxis of primary foreskin fibroblasts and in other cell types. PDGF-BB stimulation was found to lead to activation of JNK with a maximum after 30 min. Inhibition of JNK reduced Ser178 phosphorylation of the focal adhesion component paxillin. Paxillin phosphorylation at this site has been shown to be involved in the dynamics of focal adhesions and consequently cell migration. Moreover, we observed localization of JNK to the actin-dense membrane ruffles induced by PDGF-BB stimulation both using immunofluorescence staining and green fluorescent protein-tagged JNK. This suggests a role for JNK at the leading edge of the cell compatible with a function in cell migration. Furthermore, we show that phosphatidylinositol 3-kinase (PI 3-kinase), which has an established role in PDGF-stimulated cell migration, is necessary for PDGF-induced activation of JNK. In conclusion, JNK is a critical component downstream of PI 3-kinase that may be involved in PDGF-stimulated chemotaxis presumably by modulating the integrity of focal adhesions by phosphorylating its components.

In vivo molecular imaging of adenoviral versus lentiviral gene therapy in two bone formation models

Regional gene therapy techniques are promising methods to enhance bone formation in large bone defects that would be difficult to treat with allograft or autograft bone stock. In this study, we compared in vivo temporal expression patterns of adenoviral- and lentiviral-mediated gene therapy in two bone formation models. Primary rat bone marrow cells (RBMC) were transduced with lentiviral or adenoviral vectors containing luciferase (Luc) or BMP-2 cDNA, or cotransduced with vectors containing Luc and bone morphogenetic protein 2 (BMP-2). In vitro protein production was determined with luciferase assay or ELISA (for BMP-2 production) weekly for 12 weeks. Two bone formation models were used -- a hind limb muscle pouch or radial defect -- in SCID mice. A cooled charged-coupled device (CCD) camera was used to image in vivo luciferase expression weekly for 12 weeks. In vitro, adenoviral expression of BMP-2 and luciferase was detected by ELISA or luciferase assay, respectively, for 4 weeks. Lentiviral expression of BMP-2 and luciferase was sustained in culture for 3 months. Using the CCD camera, we found that adenoviral vectors expressed luciferase expression for up to 21 days, but lentiviral vectors expressed target gene expression for 3 months in vivo in both bone formation models. There was no detectable difference in the amount of bone formed between the adenoviral and lentiviral groups. Lentiviral-mediated delivery of BMP-2 can induce long term in vitro and in vivo gene expression, which may be beneficial when developing tissue engineering strategies to heal large bone defects or defects with a compromised biologic environment.

Convective Ionospheric Storms: A Major Space Weather Problem

Convective Ionospheric Storms: A Major Space Weather Problem

Optical flow visualization system for the cryogenic environment

Low temperature optical visualization provides an essential tool in understanding some types of fluid flow. We outline the various optical systems we have successfully used to visualize Rayleigh–Bénard convection in normal He4, with comparisons of image quality between each method. The current system uses a cooled charge coupled device camera at liquid nitrogen temperatures and a 1 mW laser diode light source for illumination.

Factors Influencing Quantification of In Vivo Bioluminescence Imaging: Application to Assessment of Pancreatic Islet Transplants

The aim of this study is to determine and characterize factors influencing in vivo bioluminescence imaging (BLI) and apply them to the specific application of imaging transplanted pancreatic islets. Noninvasive quantitative assessment of transplanted pancreatic islets poses a formidable challenge. Murine pancreatic islets expressing firefly luciferase were transplanted under the renal capsule or into the portal vein of nonobese diabetic-severe combined immunodeficiency mice and the bioluminescence was quantified with a cooled charge coupled device camera and digital photon image analysis. The important, but often neglected, effects of wound healing, mouse positioning, and transplantation site on bioluminescence measurements were investigated by imaging a constant emission, isotropic light-emitting bead (lambda = 600) implanted at the renal or hepatic site. The renal beads emitted nearly four times more light than hepatic beads with a smaller spot size, indicating that light absorption and scatter are greatly influenced by the transplant site and must be accounted for in BLI measurements. Detected luminescence decreased with increasing angle between the mouse surface normal and optical axis. By defining imaging parameters such as postsurgical effects, animal positioning, and light attenuation as a function of transplant site, this study develops BLI as a useful imaging modality for quantitative assessment of islets post-transplantation.