Evaluation of the molecular recognition of peptide-conjugated polymer.

We have recently reported on dodecamer peptides (HPPMDFHKAMTR, CHPQPLKSRNPL) which recognize 52-58th and 197-203rd amino acid sequences of glucose oxidase (GOx) by screening via a phage random peptide library. In this study, a side-chain protected peptide monomer (PPM) was synthesized using two peptides (HPPMDFHKAMTR, SHPQPLKSRNPL) and acryloyl chloride. The peptide-conjugated polymer (PCP) was copolymerized with PPMs and N,N-diethylacrylamide (DEAA). The affinity of PCPs to GOx was estimated using surface plasmon resonance detection. This study suggests that PCP is a valuable molecular recognition biomolecule.     

Self-assembled DNA-conjugated polymer for novel DNA chip.

We developed DNA-conjugated polymer for DNA chip fabrication. A 30 mer probe DNA and disulfide bridges were covalently attached to the polymer side chain. The DNA-conjugated polymer can be specifically adsorbed on a gold substrate surface by a self-assembly technique. The interaction between fully matched DNA and DNA-conjugated polymer was investigated by surface plasmon resonance (SPR) technique. The DNA-conjugated polymer-modified gold surface highly recognized fully matched DNA, rather than unmatched DNA. Therefore, DNA-conjugated polymer can be used for novel DNA chip fabrication.     

Analysis of tetracycline antibiotics using HPLC with pulsed amperometric detection.

The analysis of tetracycline, oxytetracyline, chlortetracycline and doxycycline by high-performance liquid chromatography with pulsed amperometric detection using an anodized boron-doped diamond thin film (BDD) electrode is originally reported. The analyses were carried out using the mobile phase, phosphate buffer (0.01 M, pH 2.5)-acetonitrile (80:20; v/v), on a C18 column (250 x 4.6 mm i.d., 5 microm) at a flow rate of 1.0 mL/min. The optimal PAD waveform parameters at the anodized BDD were 1.5 V (versus Ag/AgCl) detection potential (E(det)) for 290 ms (200 ms delay time and 90 ms integration time), 2.0 V (versus Ag/AgCl) oxidation potential (E(oxd)) for 200 ms oxidation time (t(oxd)) and 0.4 V (versus Ag/AgCI) reduction potential (E(red)) for 200 ms reduction time (t(red)). The proposed method showed the simultaneous determination of tetracycline, oxytetracyline, chlortetracycline and doxycycline with a linear range of 0.1 - 100 microg/mL, detection limits of 0.05 - 0.1 microg/mL and recoveries of 70.8 - 96.0%. The application of this method to real samples was demonstrated and validated using a shrimp sample.     

Lightness Change as Perceived in Relation to the Size of Recognized Visual Space of Illumination

According to the concept of the recognized visual space of illumination (RVSI) the lightness of an object surface is perceived in relation to its conceptualized size. To prove this proposition the lightness of gray test patches was judged when they were located at various positions inside an illuminated space composed of two rooms in the depth direction from a subject. No retinal image arrangement was changed in the test patch and its immediate surroundings, but the front room had walls, floors and furniture lower in lightness by the amount of N1.5 than the back room to make the RVSI of the former smaller despite the illuminance in the entire space being the same. The results showed that the apparent lightness of the patches was perceived higher by amount of about 13 in L units for the N4 test patch and about 20 for N6 when the patches were located in the front room, in accordance with the prediction. It was stressed that the experiment of lightness judgment should be conducted in a three dimensional space rather than two dimensional plane as done by several investigators.     

Tumor cell detection method using complement-mediated cytolytic reaction and imaging sensor system

A novel tumor-detection system consisting of complementmediated cytolytic reaction and an image processing system was developed for the simple and rapid determination of tumor cells. The present system consists of a CCD image sensor, image memory board, personal computer, and microscope. When monoclonal antibody 3C4, which is specific to the guinea pig hepatoma L-10, was added to cell suspension, only L-10 cytolysis occurred. Cytolysis caused a decrease in brightness of the cells observed by phase-contrast microscopy. The phase contrast image of the cells before cytolysis was converted to a digitalized signal and stored in computer memory. After cytolysis, a brightness threshold above that of lysed cells was subtracted from the digitalized signal and compared to the signal stored before reaction. L-10 cells in mixed cell suspension were determined specifically by the system. Measurement time was only 2 sec and overall time, including reaction time, was approximately 30 min. Since this method does not require a cell washing process, automation of the whole system is possible.     

Frequency measuring system using mirror gap stabilized Fabry-Perot interferometer

The mirror gap of a Fabry-Perot interferometer was stabilized with two laser diodes; one locked to the line frequency 385,243,555.14445 MHz of the F=3<--1 in 5D(3/2)<--5S(1/2) (87Rb) and the other to the 385,284,566.3663 MHz of the F=4<--2 in 5D(5/2)<--5S(1/2) (87Rb) [Opt. Commun. 102 (1993) 432]. The length of the mirror gap was adjusted to generate the zero-cross points simultaneously at both of the two line positions. The fringe signals obtained from an interferometer thus stabilized can be used as frequency markers having accuracies of the order of 10(10) depending on the finesse of the interferometer used. Based on measurement using Cs D1 hyperfine lines reported by Udem et al. [Phys. Rev. Lett. 82 (1999) 3568], the uncertainty of markers in the region 50 THz apart from the reference lines is +/- 5 MHz.     

Cation binding properties of crown ether-modified polyethylenimines

Crown ether-modified polyethylenimines (PEI-CR) were prepared to elucidate their cation binding efficiency and selectivity by liquid-liquid and solid-liquid extraction methods. With alkali metal cations it was found that polyethylenimines containing benzo-15-crown-5 moieties (Ia and Ib) extracted all cations examined more efficiently than their monomeric analogs, especially in the cases of K⁺ and Rb⁺ cations. The solid-liquid extraction revealed clearly that the polymers complexed with K⁺ and Rb⁺ cations according to a 1:2 cation-to-crown stoichiometry. The high selectivities of the polymers for K⁺ and Rb⁺ were interpreted in terms of the favourable conformation of the polymer chain for the formation of intramolecular 1:2 complexes. On the contrary, these polymers bound alkali earth cations less effectively with low selectivities.     

Vibrational spectra and molecular conformations of 1,5-dichloro- and 1,5-dibromopentanes and 1,6-dichloro-and 1,6-dibromohexanes

The Raman and IR spectra of 1,5-dichloro- and 1,5-dibromopentanes and 1,6-dichloro-and 1,6-dibromohexanes have been measured. The normal coordinates have been calculated for these molecules using a consistent set of force constants and the molecular conformations studied by analysing the spectra with reference to the results of the calculations. In the crystalline solid state, 1,5-dichloropentane assumes the trans-trans-trans-gauche form and 1,5-dibromopentane, 1,6-dichloro- and 1,6-dibromohexanes assume the all-trans form. The normal coordinate treatment with the well-established force field was of great help in determining the whole molecular form of the relatively large chain molecules.     

Fluorimetric assay for measuring Dns-His-Lys-Arg-His-Lys cleaving enzyme using high-performance liquid chromatography

A rapid and sensitive assay for the determination of Dns-His-Lys-Arg-His-Lys cleaving enzyme activity is reported. This assay is based on fluorimetric detection of a dansylated dipeptide, 5-dimethylaminonaphthalene-1-sulfonyl-His-Lys, enzymatically formed from the substrate 5-dimethylaminonaphthalene-1-sulfonyl-His-Lys-Arg-His-d-Lys, after separation by high-performance liquid chromatography (HPLC) using a C-18 reversed-phase column by isocratic elution. This assay is sensitive enough to measure 5-dimethylaminonaphthalene-1-sulfonyl-His-Lys at concentrations as low as 7 pmol, and yields highly reproducible results and requires less than 9.0 min per sample for separation and quantitation. The optimum pH for Dns-His-Lys-Arg-His-Lys cleaving enzyme activity was 7.5-8.0. The Michaelis constant (K_m) and the maximum velocity (V_max) values were 33.3 μM and 47.07 pmol/(μg h), respectively with the use of enzyme extract obtained from bovine pituitary. By using this assay, axonal transport of this enzyme activity was observed 48 h after double ligations of rat sciatic nerves. The high sensitivity and selectivity of this assay would be useful for clarification of the physiological role of this enzyme.