A novel biofunctionalizing peptide for metallic alloy.

Objectives: Improving biocompatibility of metallic alloy biomaterials has been of great interest to prevent implant associated-diseases, such as stent thrombosis. Herein a simple and efficient procedure was designed to biofunctionalize a biomaterial surface by isolating a SUS316L stainless steel binding peptide.

Results: After three rounds of phage panning procedure, 12 mer peptide (SBP-A; VQHNTKYSVVIR) was identified as SUS316L-binding peptide.

Stabilization of G-quadruplex structure on vascular endothelial growth factor gene promoter depends on CpG methylation site and cation type.

Background: DNA methylation at the 5-position of cytosine is an epigenetic modification of CpG dinucleotides. In addition to CpG methylation, the G-quadruplex (G4) structure has been reported as a regulator of gene expression. The identification of G4 forming sequences in CpG islands suggests an involvement of CpG-methylated G4 structures in biological processes; however, few reports have addressed the effects of CpG methylation on G4 structure.

Identification of G-quadruplex clusters by high-throughput sequencing of whole-genome amplified products with a G-quadruplex ligand.

G-quadruplex (G4) is a DNA secondary structure that has been found to play regulatory roles in the genome. The identification of G4-forming sequences is important to study the specific structure-function relationships of such regions. In the present study, we developed a method for identification of G4 clusters on genomic DNA by high-throughput sequencing of genomic DNA amplified via whole-genome amplification (WGA) in the presence of a G4 ligand.

A quantitative homogeneous assay for global DNA methylation levels using CpG-binding domain- and methyl-CpG-binding domain-fused luciferase.

Global DNA methylation levels have been considered as biomarkers for cancer diagnostics because transposable elements that constitute approximately 45% of the human genome are hypomethylated in cancer cells. We have previously reported a homogeneous assay for measuring methylated CpG content of genomic DNA based on bioluminescence resonance energy transfer (BRET) using methyl-CpG-binding domain (MBD)-fused luciferase (MBD-luciferase). In this study, a homogeneous assay for measuring unmethylated CpG content of genomic DNA in the same platform was developed using CXXC domain-fused luciferase (CXXC-luciferase) that specifically recognizes unmethylated CpG.

Identification of G-quadruplex structures that possess transcriptional regulating functions in the Dele and Cdc6 CpG islands.

Background: G-quadruplex is a DNA secondary structure that has been shown to play an important role in biological systems. In a previous study, we identified 1998 G-quadruplex-forming sequences using a mouse CpG islands DNA microarray with a fluorescent-labeled G-quadruplex ligand. Among these putative G-quadruplex-forming sequences, G-quadruplex formation was verified for 10 randomly selected sequences by CD spectroscopy and DMS footprinting analysis.

Global DNA Methylation Detection System Using MBD-Fused Luciferase Based on Bioluminescence Resonance Energy Transfer Assay.

DNA methylation plays an important role in the regulation of gene expression. In normal cells, transposable elements that constitute approximately 45% of the human genome are highly methylated to silence their expression. In cancer cells, transposable elements are hypomethylated; therefore, global DNA methylation level is considered as a biomarker for cancer diagnostics.

Detection of DNA Methylation of G-Quadruplex and i-Motif-Forming Sequences by Measuring the Initial Elongation Efficiency of Polymerase Chain Reaction.

DNA methylation has been proposed as one of the promising biomarkers for cancer diagnosis. In this study, we developed a DNA methylation detection system utilizing G-quadruplex and i-motif-forming sequences that requires neither sodium bisulfite treatment nor methylated DNA ligands. We hypothesized that G-quadruplex and i-motif structures would be stabilized by DNA methylation and arrest DNA polymerase activity during quantitative polymerase chain reaction (qPCR).

Sensitive detection of ochratoxin A in wine and cereals using fluorescence-based immunosensing.

Ochratoxin A (OTA) is a mycotoxin found in a wide range of food and feedstuffs. Intake of OTA-contaminated food causes health concern due to the harmful effects reported on humans and animals. Much effort is currently devoted to set up and optimise highly sensitive and accurate methods of OTA analysis.

Development of urinary 8-hydroxy-2'-deoxyguanosine (8-OHdG) measurement method combined with SPE.

A novel separation method of urinary 8-hydroxy-2'-deoxyguanosine (8-OHdG) was developed. The C(18) and the strong cation-exchange (SCX) columns were employed to separate urinary 8-OHdG. The major interfering substances were removed by the consecutive processes of the C(18) and the SCX columns.

Determination of human serum 8-hydroxy-2'-deoxyguanosine (8-OHdG) by HPLC-ECD combined with solid phase extraction (SPE).

Human serum 8-hydroxy-2'-deoxyguanosine (8-OHdG) was measured by HPLC-ECD method combined with solid phase extraction (SPE) developed by our group: (our proprietary kit, named 8-OHdG Pre-treatment Kit (TANITA Corporation)). The major interfering substances and proteins in serum were removed by 8-OHdG Pre-treatment Kit. This measurement method was highly reproducible (CV=2.

Yeast based sensors.

Since the first microbial cell sensor was studied by Karube et al. in 1977, many types of yeast based sensors have been developed as analytical tools. Yeasts are known as facultative anaerobes.

Applications of microbial cell sensors.

Since the first microbial cell sensor was studied by Karube et al. in 1977, many types of microbial cell sensors have been developed as analytical tools. The microbial cell sensor utilizes microbes as a sensing element and a transducer.

High-sensitive flow-based kinetic exclusion assay for okadaic acid assessment in shellfish samples.

Episodes of shellfish contamination with okadaic acid (OA) are a human health threat that is causing increasing concern. As a way to overcome the shortcomings involved in the reference methods of analysis set by legislations, alternative procedures are envisaged. This paper describes the development of different immunosensors for the analysis of OA, focusing on the comparison of their sensitivity, precision, ease of use and sample matrix effects.

Novel fluorescent probe for analysis of hydroperoxides based on boron dipyrromethane fluorophore.

A new reagent, N-[2-(diphenylphosphino)ethyl]-4-(1,3,5,7-tetramethyl-4,4-difluoro-4-bora-3a,4a-diaza-s-indacene-8-yl)benzamide (DPPEA-BODIPY), was designed and synthesized for analysis of hydroperoxides. DPPEA-BODIPY fluoresces at low levels in the visible range (lambda(ex)/lambda(em) = 502 nm/515 nm) and reacts with hydroperoxides to produce DPPEA-BODIPY oxide, which fluoresces at high levels. The fluorescence intensity of the reaction mixture was observed to be linearly related to the methyl linoleate hydroperoxide (MeLOOH) concentration.

Novel automated pulse immunoassay for human alpha-fetoprotein.

Background: To improve current alpha-fetoprotein (AFP) assays, which are expensive and time-consuming, a specific AFP reagent has been developed for practical use in our newly developed high-speed, highly sensitive pulse immunoassay (PIA) system, in which a latex immunoagglutination reaction is carried out under a high-frequency pulse voltage, leading to an enhanced immunological reaction.

Methods: We evaluated the assay performance (reproducibility, sensitivity, dilution linearity, interference) of the newly developed automated AFP PIA compared with the current AFP assay.

Results: Using pooled serum samples, the within-run reproducibility resulted in a correlation variation of 3.

Flow injection microbial trichloroethylene sensor.

A flow type microbial biosensor for direct measurement of trichloroethylene (TCE) was developed. The unique features of this novel microbial sensor were the use of the TCE degrading bacterium Pseudomonas aeruginosa JI104, the electrical detection of the chloride ion released by microbial degradation, and flow cell made of glass. Glass cell was used in order to suppress adsorption of TCE and made a closed reaction cell.

Effect of the polarization and incident angle of excitation light on the fluorescence enhancement observed with a multilayered substrate fabricated by Ag and Al2O3.

The fluorescence from a fluorophore on a multilayered substrate fabricated by a metal and a dielectric is known to be enhanced by more than 100-fold. In the course of this study, we prepared a multilayered substrate with Ag as the metal and Al(2)O(3) as the dielectric and then investigated the effects of the polarization of the excitation light on the enhancement of the multilayered substrate. It was found that the enhancement was attributed to an electric field oscillating parallel to the substrate.

Absorption-based highly sensitive and reproducible biochemical oxygen demand measurement method for seawater using salt-tolerant yeast Saccharomyces cerevisiae ARIF KD-003.

Salt-tolerant yeast Saccharomyces cerevisiae ARIF KD-003 was applied to highly sensitive and reproducible absorbance-based biochemical oxygen demand (BOD(AB-ScII)) measurement for seawater. In the previous work, we have studied the BOD(AB-ScI) method using normal Baker's yeast S. cerevisiae, and the excellent feature of the Baker's yeast as uniformly sustainable in solution could successfully be utilized.

An enzyme-chromogenic surface plasmon resonance biosensor probe for hydrogen peroxide determination using a modified Trinder's reagent.

An absorption-based surface plasmon resonance (SPR(Abs)) biosensor probe has been developed for simple and reproducible measurements of hydrogen peroxide using a modified Trinder's reagent (a chromogenic reagent). The reagent enabled the determination of the hydrogen peroxide concentration by the development of deep color dyes (lambda(max)=630 nm) through the oxidative coupling reaction with N-ethyl-N-(2-hydroxy-3-sulfopropyl)-3,5-dimethylaniline sodium salt monohydrate (MAOS; C(13)H(20)NNaO(4)S.H(2)O) and 4-aminoantipyrine (4-AA) in the presence of hydrogen peroxide and horseradish peroxidase (HRP).

A surface plasmon resonance probe without optical fibers as a portable sensing device.

A surface plasmon resonance (SPR) sensor integrating a small sensor probe, a laser emission diode, a photo detector, and a polarizer was developed as a portable sensing device. The sensor probe was made with a glass cylinder, 50mm long and 1.5mm in diameter, that was connected directly to a beam splitter without optical fibers.

A new diagnostic method for soil-borne disease using a microbial biosensor.

We have developed a new system for the rapid diagnosis of soil-borne diseases, consisting of two biosensors. The system was constructed using equal quantities of two different microbes, each individually immobilized on an electrode. These two sensors were coupled as a dual sensor system and used for simultaneous measurements.

SNPs typing based on the formation of fluorescent signaling DNA aptamers which bind to bile acids.

A simple and rapid method for the detection of single nucleotide polymorphisms (SNPs) is essential for the development of personalized medicine because SNPs correlate with some diseases and side effects of some drugs. Here we report a new method for the fluorescent detection of single nucleotide mutations that is based on the formation of cholic-acid binding DNA aptamers which form fully matched three-way junctions.

Development of microbial sensors and their application.

Many types of microbial sensors have been developed as analytical tools since the first microbial sensor was studied by Karube et al. in 1977. The microbial sensor consists of a transducer and microbe as a sensing element.