In vivo Preclinical Tumor-Specific Imaging of Superparamagnetic Iron Oxide Nanoparticles Using Magnetic Particle Imaging for Cancer Diagnosis.

Purpose: Magnetic particle imaging (MPI) is an emerging radiation-free, non-invasive three-dimensional tomographic technology that can visualize the concentrations of superparamagnetic iron oxide nanoparticles (SPIONs). To verify the applicability of the previously proposed point-of-care testing MPI (PoCT-MPI) in medical diagnosis and therapeutics, we imaged SPIONs in animal tumor models. Methods: CT26 or MC38 mouse colon carcinoma cells (2 × 106 cells) were subcutaneously injected into the right flank of BALB/c mice. SPIONs were either injected directly into the tumor lesions in the intratumoral group or through tail veins in the intravenous group. CT26 and MC38 tumor models were examined both intratumorally and intravenously to confirm the biological availability of SPIONs using PoCT-MPI. Results: Signals were observed in the tumor lesions from day 1 to day 7. This is the first study to successfully image the pathological region and show the biodistribution of SPIONs in CT26 tumor models using the recently developed PoCT-MPI technology. Furthermore, MC38 tumor models were examined, resulting in similar images to those of the CT26 tumor model in both intratumoral and intravenous groups. Conclusion: The present study demonstrates the biological applicability of PoCT-MPI, which promises to be a powerful diagnostic and therapeutic technique in biomedical imaging.

A novel three-dimensional magnetic particle imaging system based on the frequency mixing for the point-of-care diagnostics.

The magnetic particle imaging (MPI) is a technology that can image the concentrations of the superparamagnetic iron oxide nanoparticles (SPIONs) which can be used in biomedical diagnostics and therapeutics as non-radioactive tracers. We proposed a point-of-care testing MPI system (PoCT-MPI) that can be used for preclinical use for imaging small rodents (mice) injected with SPIONs not only in laboratories, but also at emergency sites far from laboratories. In particular, we applied a frequency mixing magnetic detection method to the PoCT-MPI, and proposed a hybrid field free line generator to reduce the power consumption, size and weight of the system. The PoCT-MPI is [Formula: see text] in size and weighs less than 100 kg. It can image a three-dimensional distribution of SPIONs injected into a biosample with less than 120 Wh of power consumption. Its detection limit is [Formula: see text], 10 mg/mL, [Formula: see text] (Fe).

Construction of 3D-rendering imaging of an ischemic rat brain model using the planar FMMD technique.

Occlusion of the major cerebral artery usually results in brain hypoxic-ischemic injury, which evokes neuroinflammation and microglial activation. Activated microglia are considered a source of multiple neurotoxic factors, such as reactive oxygen species (ROS), in the central nervous system (CNS). We herein present a 3D-rendering brain imaging technique in an experimental rodent model of cerebral ischemia based on 2D magnetic images of superparamagnetic iron oxide nanoparticles (SPIONs) using the planar frequency mixing magnetic detection (p-FMMD) technique. A rat model of cerebral ischemia was established by unilateral middle cerebral artery occlusion with reperfusion (MCAO/R) injury. 2,3,5-Triphenyltetrazolium chloride (TTC) staining was performed to demonstrate the irreversibly damaged ischemic brain tissues, and double immunofluorescent labeling of OX6 (activated microglial marker) and ethidium (ROS marker) was conducted to confirm ROS generation in the activated microglia in the infarcted brain region. The ischemic brain sections treated with OX6-conjugated SPIONs were scanned using our p-FMMD system, yielding 2D images on the basis of the nonlinear magnetic characteristics inherent in SPIONs. The p-FMMD signal images representing microglia activation show an infarct ratio of 44.6 ± 7.1% compared to the contralateral counterpart, which is smaller than observed by TTC (60.9 ± 4.9%) or magnetic resonance imaging (MRI, 65.7 ± 2.7%). Furthermore, we developed a 3D-rendering brain imaging process based on the 2D p-FMMD signal images. The 3D reconstructed model showed a decreased ratio of coincidence of the ischemic regions compared with MRI models. In this study, we successfully conducted a feasibility test on whether our p-FMMD technology, a technique for signaling and imaging based on the nonlinearity of SPIONs, can be used to visualize the ischemic brain region in real time by detecting activated microglia in an MCAO/R animal model. Therefore, our method might allow for a different approach to analyze the pathophysiology of ischemic stroke through molecular imaging. Furthermore, we propose that this magnetic particle imaging (MPI) technique that detects the nonlinear magnetization properties of SPIONs could be applied not only to a stroke model but also to various types of pathophysiological studies as a new bioimaging tool.

Frequency Mixing Magnetic Detection Scanner for Imaging Magnetic Particles in Planar Samples.

The setup of a planar Frequency Mixing Magnetic Detection (p-FMMD) scanner for performing Magnetic Particles Imaging (MPI) of flat samples is presented. It consists of two magnetic measurement heads on both sides of the sample mounted on the legs of a u-shaped support. The sample is locally exposed to a magnetic excitation field consisting of two distinct frequencies, a stronger component at about 77 kHz and a weaker field at 61 Hz. The nonlinear magnetization characteristics of superparamagnetic particles give rise to the generation of intermodulation products. A selected sum-frequency component of the high and low frequency magnetic field incident on the magnetically nonlinear particles is recorded by a demodulation electronics. In contrast to a conventional MPI scanner, p-FMMD does not require the application of a strong magnetic field to the whole sample because mixing of the two frequencies occurs locally. Thus, the lateral dimensions of the sample are just limited by the scanning range and the supports. However, the sample height determines the spatial resolution. In the current setup it is limited to 2 mm. As examples, we present two 20 mm × 25 mm p-FMMD images acquired from samples with 1 µm diameter maghemite particles in silanol matrix and with 50 nm magnetite particles in aminosilane matrix. The results show that the novel MPI scanner can be applied for analysis of thin biological samples and for medical diagnostic purposes.

Magnetic immunoassay platform based on the planar frequency mixing magnetic technique.

We represent the experimental results of our planar-frequency mixing magnetic detection (p-FMMD) technique to obtain 2D superparamagnetic images for magnetic immunoassay purpose. The imaging of magnetic beads is based on the nonlinear magnetic characteristics inherent in superparamagnetic materials. The p-FMMD records the sum-frequency components originating from both a high and a low frequency magnetic field incident on the magnetically nonlinear nanoparticles. In this study, we apply the p-FMMD technique to 2D scanning imaging of superparamagnetic iron oxide nanoparticles (SPIONs) in a microfluidic platform. Our p-FMMD system enables to acquire planar images of SPIONs filled in a microchannel as narrow as 30µm in width. The minimum detectable amount is ~1.0×10(8) beads of 100nm size. The system shows a spatial resolution enabling to distinguish between two distinct channels even 2mm apart from each other. Our p-FMMD system as a magnetic immunoassaying system has permitted the detection of amyloid beta 42 (Aß42), a promising biomarker of Alzheimer's disease, at the minimum concentration of 23.8pg/ml. This may enable the identification of the Aß42 levels for the early-stage of Alzheimer's disease with the assistance of the MPI using p-FMMD technique. The results show that the deployment of the p-FMMD can be an alternative to conventional biosensing analytical methods, and can be used as a fast and portable screening method.

Optoelectronic Characterization of Infrared Photodetector Fabricated on Ge-on-Si Substrate.

We report on the optoelectronic characterization of Ge p-i-n infrared photodetector fabricated on Ge-on-Si substrate using rapid thermal chemical vapor deposition (RTCVD). The phosphorous doping concentration and the root mean square (RMS) surface roughness of epitaxial layer was estimated to be 2 x 10(18) cm(-3) and 1.2 nm, respectively. The photodetector were characterized with respect to their dark, photocurrent and responsivities in the wavelength range of 1530-1630 nm. At 1550 nm wavelength, responsivity of 0.32 A/W was measured for a reverse bias of 1 V, corresponding to 25% external quantum efficiency, without an optimal antireflection coating. Responsivity drastically reduced from 1560 nm wavelength which could be attributed to decreased absorption of Ge at room temperature.

Magnetic particle imaging with a planar frequency mixing magnetic detection scanner.

We present the first experimental results of our planar-Frequency Mixing Magnetic Detection (p-FMMD) technique to obtain Magnetic Particles Imaging (MPI). The p-FMMD scanner consists of two magnetic measurement heads with intermediate space for the analysis of the sample. The magnetic signal originates from the nonlinear magnetization characteristics of superparamagnetic particles as in case of the usual MPI scanner. However, the detection principle is different. Standard MPI records the higher order harmonic response of particles at a field-free point or line. By contrast, FMMD records a sum-frequency component generated from both a high and a low frequency magnetic field incident on the magnetically nonlinear particles. As compared to conventional MPI scanner, there is no limit on the lateral dimensions of the sample; just the sample height is limited to 2 mm. In addition, the technique does not require a strong magnetic field or gradient because of the mixing of the two different frequencies. In this study, we acquired an 18 mm × 18 mm image of a string sample decorated with 100 nm diameter magnetic particles, using the p-FMMD technique. The results showed that it is feasible to use this novel MPI scanner for biological analysis and medical diagnostic purposes.

Is the biotransformation of chlorinated dibenzo-p-dioxins by Sphingomonas wittichii RW1 governed by thermodynamic factors?

Density functional theory (DFT) calculations were used to explore the relationship between the biotransformation of dibenzo-p-dioxin and selected chlorinated derivatives by resting cells of Sphingomonas wittichii RW1 and measuring the thermodynamic properties of the biotransformation substrates. Sphingomonas wittichii RW1 can aerobically catabolize dibenzo-p-dioxin as well as 2,7-dichloro-, 1,2,3-trichloro-, 1,2,3,4-tetrachloro-, and 1,2,3,4,7,8-hexachlorodibenzo-p-dioxin; however, neither the 2,3,7-trichloro- nor the 1,2,3,7,8-pentachlorodibenzo-p-dioxin was transformed to its corresponding metabolic intermediate. The experimental biotransformation rates established were apparently governed by the selected thermodynamic properties of the substrates tested.

In situ measurement of superoxide and hydroxyl radicals by frequency mixing detection technique.

Frequency mixing magnetic detection (FMMD) was used to detect superoxide from hypoxanthine and xanthine reaction and to detect hydroxyl radical from the Fenton reaction. FMMD was also applied to measure the reactive oxygen species (ROS) level released from microglial cells. We could assess the formation and extinction of the free radicals without a spin trap reagent. The FMMD signal amplitude scaled with the concentration of the radicals. It was verified that no signals are obtained from the substrates and reagents. Based on the observations and on previous research, we suggest that the FMMD signals originate from superoxide and hydroxyl radicals, indicating that FMMD can be used to detect O-centered radicals. Subsequent analysis of free radicals generated from living microglial cells showed that there were significant differences between the activated microglial cells and resting ones. The results of this research are promising regarding the applications of FMMD for in situ measurement of free radicals from various sources, including the cell.

Analysis of beta amyloid peptides employing the small probe molecules.

We herein describe an analytical method employing a small molecule array for the characterization of similar proteins based on ligand binding. In this study, 2 different beta amyloids (Abeta(1-40) and (1-42)) were selected as the model compounds. Their primary structures are identical except for 2 additional C-terminal amino acids. However, many studies have observed different biological and chemical characteristics of these peptides. Thus, the ability to distinguish these 2 peptides is important in the diagnosis and development of treatments for related disorders such as Alzheimer's disease. However, strong non-specific binding is usually observed, even when specific antibodies for each peptide are employed. In this study, Abeta(1-40) and Abeta(1-42) peptides were immobilized on a typical 96-well microplate. Twenty different small probe molecules (modified amino acids conjugated with FITC) were applied to the peptides acting as the secondary antibodies and labeling compounds. The results show that specific binding patterns occurred according to Abeta type and the analysis of the patterns can be used to distinguish these 2 similar peptides.

Detection of two different influenza A viruses using a nitrocellulose membrane and a magnetic biosensor.

Here we describe a new analytical method for the detection of two influenza A viruses by nitrocellulose membrane and magnetic sensors that employ a special frequency mixing technique. The combination of the nitrocellulose membrane and magnetic bead detection permits a rapid assay procedure and excludes two steps (the development of color and the stop reaction) required for usual immunochemical detection methods such as ELISA. Quantitative virus detection was performed using magnetic beads conjugated with secondary antibody. The results were compared with conventional assay methods and with a dot-blot assay with fluorescence compound (FITC). Under optimum conditions, our new assay procedure is capable of detecting picograms of virus per well. This new method combining the nitrocellulose membrane and magnetic bead detection reduces analytical time and allows stable and repeatable analyses of samples in point-of-care applications.

Effect of heavy metals on the biodegradation of dibenzofuran in liquid medium.

The effect of heavy metals on the degradation of dibenzofuran by Sphingomonas wittichii RW1 was determined in liquid cultures. The results showed that 10mg/L cadmium, mercury and copper not only affected the growth of RW1 with dibenzofuran but also the ability of resting cells to degrade this compound. Growth and degradation were strongly inhibited by mercury, even at 1mg/L, while the inhibitory effect of cadmium and copper at the same concentration or at 5mg/L were negligible. In contrast, arsenic and lead did not affect degradation or growth, even at very high concentrations of 100mg/L. Subsequent analyses additionally revealed that concentrations of arsenic and lead remained unchanged following incubation, while those of cadmium, mercury and copper decreased significantly.

Biological removal of polychlorinated dibenzo-p-dioxins from incinerator fly ash by Sphingomonas wittichii RW1.

The ability of Sphingomonas wittichii strain RW1 to remove polychlorinated dibenzo-p-dioxins (PCDDs) from fly ash was investigated. All experiments were carried out in a slurry-phase system. Preliminary studies with resting cells of strain RW1 in a model fly ash system showed the complete removal of dibenzofuran (DF) and 81% of dibenzo-p-dioxin (DD). Incubation of real fly ash collected from municipal waste incinerators with strain RW1 for 15 days resulted in a 75.5% reduction in toxic PCDDs. When the same experiment was carried out using dead strain RW1 cells a 20.2% reduction in toxic PCDDs was observed, indicating that adsorption onto biomass was an important factor in dioxin elimination. Further analyses revealed that live strain RW1 cells removed 83.8% of the 2,3,7,8-substituted congeners from the fly ash, while dead cells removed 32.1% of the same congeners. To enhance the removal efficiency of toxic PCDDs, the effects of adding surfactant, repeated inoculation, and pre-adaptation of cultures were also studied. The removal of toxic PCDDs was enhanced by up to 10.3% upon repeated inoculation of the strain RW1, but was not much affected by the addition of surfactant. The present results suggest that S. wittichii strain RW1 is a potential candidate for the industrial removal of PCDDs from incinerator fly ash.

Biodegradation of dibenzo-p-dioxin, dibenzofuran, and chlorodibenzo-p-dioxins by Pseudomonas veronii PH-03.

The dioxin-degrading strain Pseudomonas veronii PH-03 was isolated from contaminated soil by selective enrichment techniques. Strain PH-03 grew on dibenzo-p-dioxin and dibenzofuran as a sole carbon source. Further, 1-chlorodibenzo-p-dioxin, 2-chlorodibenzo-p-dioxin and other dioxin metabolites, salicylic acid, and catechol were also metabolized well. Resting cells of strain PH-03 transformed dibenzo-p-dioxin, dibenzofuran, 2,2',3-trihydroxybiphenyl, and some chlorodioxins to their corresponding metabolic intermediates such as catechol, salicylic acid, 2-hydroxy-(2-hydroxyphenoxy)-6-oxo-2,4-hexadienoic acid, and chlorocatechols. The formation of these metabolites was confirmed by comparison of gas chromatography-mass spectrometry (GC-MS) data with those of authentic compounds. Although we did observe the production of 3,4,5,6-tetrachlorocatechol (3,4,5,6-TECC) from 1,2,3,4-tetrachlorodibenzo-p-dioxin (1,2,3,4-TCDD) with resting cell suspensions of PH-03, growth of strain PH-03 in the presence of 1,2,3,4-TCDD was poor. This result suggests that strain PH-03 is unable to utilize 3,4,5,6-TECC, even at very low concentration (0.01 mM) due to its toxicity. In cell-free extracts of DF-grown cells, 2,2',3-trihydroxybiphenyl dioxygenase, 2-hydroxy-6-oxo-6-phenyl-2,4-hexadienoic acid hydrolase, and catechol-2,3-dioxygense activities were detected. Moreover, the activities of meta-pyrocatechase and 2,2',3-trihydroxybiphenyl dioxygenase from the crude cell-free extracts were inhibited by 3-chlorocatechol. However, no inhibition was observed in intact cells when 3-chlorocatechol was formed as intermediate.

Adsorption of halogenated aromatic pollutants by a protein released from Bacillus pumilus.

Previous studies of the biosorption of halogenated aromatic pollutants (HAPs) have focused on the sorption of these compounds by cell bodies. However, in this study we investigated the adsorption of HAPs by biocompounds released from a bacterium, Bacillus pumilus. When B. pumilus was exposed to high temperature, it released a protein and carbohydrates, exclusively. After determining experimental conditions using 1,2,3,4-tetrachlorinated dibenzofuran (1,2,3,4-TCDF), the adsorption characteristics of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs), polychlorinated biphenyls (PCBs), chlorinated benzenes, and chlorinated naphthalenes were investigated. These HAPs were adsorbed considerably not only by cells but also by the released protein. In general, highly chlorinated congeners were adsorbed to a greater extent on the protein than lowly chlorinated ones, and the amount adsorbed differed between isomers. The present results are consistent with adsorption occurring via a passive physico-chemical mechanism. Finally, the importance of biocompounds released or excreted from microorganisms for the removal of HAPs is discussed.

Biotransformation of 2,7-dichloro- and 1,2,3,4-tetrachlorodibenzo-p-dioxin by Sphingomonas wittichii RW1.

Aerobic biotransformation of the diaryl ethers 2,7-dichlorodibenzo-p-dioxin and 1,2,3,4-tetrachlorodibenzo-p-dioxin by the dibenzo-p-dioxin-utilizing strain Sphingomonas wittichii RW1, producing corresponding metabolites, was demonstrated for the first time. Our strain transformed 2,7-dichlorodibenzo-p-dioxin, yielding 4-chlorocatechol, and 1,2,3,4-tetrachlorodibenzo-p-dioxin, producing 3,4,5,6-tetrachlorocatechol and 2-methoxy-3,4,5,6-tetrachlorophenol; all of these compounds were unequivocally identified by mass spectrometry both before and after N,O-bis(trimethylsilyl)-trifluoroacetamide derivatization by comparison with authentic standards. Additional experiments showed that strain RW1 formed a second metabolite, 2-methoxy-3,4,5,6-tetrachlorophenol, from the original degradation product, 3,4,5,6-tetrachlorocatechol, by methylation of one of the two hydroxy substituents.