Transient cell proliferation with polyethylenimine-cationized N-terminal domain of simian virus 40 large T-antigen

Polyethylenimine (PEI) cationization is a powerful strategy for protein transduction into cells. In this study, we attempted the artificial regulation of cell proliferation by protein transduction of the N-terminal domain (1-132 amino acids) of the simian virus 40 large T-antigen (SVLT-N), which inactivates retinoblastoma family proteins but not p53. To deliver SVLT-N into cells, we employed an indirect cationization method by forming a complex of biotynylated SVLT-N through disulfide bonds (biotin-SS-SVLT-N) and PEI-cationized avidin (PEI600-avidin). Using this complex, SVLT-N was transduced into the nucleus of confluent and quiescent Balb/c 3T3 cells and was found to be complexed with a cellular target protein, pRb. Furthermore, SVLT-N transduction induced cell proliferation in spite of confluent conditions. Because SVLT-N thus transduced into cells gradually degraded and was not detectable after a 4-d incubation, transiently transformed cells were obtained by this method. These results suggest that oncogene protein transduction technology has great potential for in vitro regulation of cell proliferation.     

Characteristics of the thiobarbituric acid reactivity of oxidized fats and oils

The thiobarbituric acid (TBA) reactivity of oxidized methyl linoleate, soybean oil, sesame oil, lard, chicken oil and sardine oil was characterized by using four different methods with 0.01% butylated hydroxytoluene (BHT). Optimal pH for the reactivity of most of the oxidized samples was 3–4, and that of some samples was above 5. Introduction of 2 mMt-butyl hydroperoxide (t-Bu00H) or 0.2 mM ferric ion in the reaction markedly enhanced the reactivity. Introduction of 0.2 mM ethylenediamine tetraacetic acid suppressed the reactivity. The characteristics of the TBA-reactivity of the samples were similar to those of alkadienals or alkenals. The most preferable method for the estimation of the TBA-reactive substances of the oxidized fats and oils was that using solvents at pH 3.5 with introduction of BHT, andt-Bu00H or ferric ion.     

3D true-phase polarity recovery with independent phase estimation using three-tier stacks based region growing (3D-TRIPS)

Objectives

A postprocessing technique termed 3D true-phase polarity recovery with independent phase estimation using three-tier stacks based region growing (3D-TRIPS) was developed, which directly reconstructs phase-sensitive inversion-recovery images without acquisition of phase-reference images. The utility of this technique is demonstrated in myocardial late gadolinium enhancement (LGE) imaging.

Materials and methods

A data structure with three tiers of stacks was used for 3D-TRIPS to directly achieve reliable region growing for successful background-phase estimation. Fifteen patients undergoing postgadolinium 3D phase-sensitive inversion recovery (PSIR) cardiac LGE magnetic resonance imaging (MRI) were recruited, and 3D-TRIPS LGE reconstructions were compared with standard PSIR. Objective voxel-by-voxel comparison was performed. Additionally, blinded review by two radiologists compared scar visibility, clinical acceptability, voxel polarity error, or groups and blurring.

Results

3D-TRIPS efficiently reconstructed postcontrast phase-sensitive myocardial LGE images. Objective analysis showed an average 95% voxel-by-voxel agreement between 3D-TRIPS and PSIR images. Blinded radiologist review demonstrated similar image quality between 3D-TRIPS and PSIR reconstruction.

Conclusion

3D-TRIPS provided similar image quality to PSIR for phase-sensitive myocardial LGE MRI reconstruction. 3D-TRIPS does not require acquisition of a reference image and can therefore be used to accelerate phase-sensitive LGE imaging.

     

Does Globalization Reduce or Enhance Democracy? A Micro Analysis Based on an 18 Country Cross-National Asia-Europe Survey

Does globalization have a positive or negative impact on democracy? One reason this problem has gone unsolved is found in the fact that most studies to date have not made systematic use of empirical data to test propositions concerning the relationship between globalization and democracy. While there have been studies that have made a pioneering systematic contribution through the use of macroeconomic and other aggregate statistics, this article empirically examines whether globalization enhances or constrains democracy by using cross-national survey data collected in 17 countries (the Asia-Europe Survey). Our empirical testing has shown that globalization tends to be positively correlated to democratic activism at the individual level, suggesting the possibility that experiences of globalization strengthen democracy.

     

Noncompetitive detection of low molecular weight peptides by open sandwich immunoassay

Small peptides with less than 1000 in molecular weight are not considered amenable to sandwich immunoassays due to their difficulty of simultaneous recognition by two antibodies. As an alternative, we attempted noncompetitive detection of small peptides by open sandwich enzyme-linked immunosorbent assay (OS-ELISA) utilizing the antigen-induced enhancement of antibody VH/VL interaction. Taking fragments of human osteocalcin (BGP), a major non-collagen peptide produced in bone, as model peptides, OS immunoassay was performed using the cloned VH and VL cDNAs from two anti-BGP monoclonal antibodies either recognizing the N- or C-terminal fragment, respectively. When the clones were used for OS-ELISA with immobilized VL fragment and phage-displayed VH fragment, enhanced VH/VL interaction upon BGP addition was observed. Especially the clone for the C-terminal fragment showed a superior detection limit as well as a wider working range than those of competitive assay. The result was reproduced with purified VH-alkaline phosphatase and MBP-VL fusion proteins, where the latter was directly immobilized onto the microplate wells. The minimum detectable fragment was the hexamer including the C-terminus. This simple approach with a single monoclonal antibody with a short measurement time may prove a useful tool in immunodiagnostics as well as in proteomics research.     

A new whiteness formula

With the advent of fluorescent whitening agents, the general idea of whiteness has changed. Numerous whiteness formulas by which we evaluate whiteness of white materials have been proposed, but these formulas (including the CIE whiteness formula) were found not to correlate well to visual estimation. In this article, a modified CIE whiteness formula dealing with the tint and the excitation purity is proposed. There is a significant correlation between this formula and the visual estimation of whiteness. © 1998 John Wiley & Sons, Inc. Col Res Appl, 23, 202–209, 1998     

Simultaneous determination of pesticides in agricultural products by LC/MS/MS using clean-up with ultrafiltration

A method for simultaneous determination of multiple pesticide residues in agricultural products was developed by using a pretreatment with ultrafiltration, followed by liquid chromatography-tandem mass spectrometry (LC/MS/MS). The pretreatment process (extraction of pesticides from agricultural products with methanol, dilution of the extract with water, and ultrafiltration) gave recoveries in the range of 50-150% for 63 of 83 pesticides spiked at 0.25 microg/ g into 6 agricultural products. The detection limits of pesticides by LC/MS/MS were below 0.0005-0.05 micro/g. This method is useful for screening purposes and for multiresidue analysis of pesticides in agricultural products. Pesticide residues in 50 domestic crops were investigated by this method, and residues of 14 pesticides were detected in 30 crops.     

Solution-processed zinc tetrabenzoporphyrin thin-films and transistors

Thin-films and organic field-effect transistors fabricated from a solution-processable precursor of zinc tetrabenzoporphyrin (ZnTBP) are reported. Amorphous, insulating precursor films were deposited by spin-casting and thermally converted into polycrystalline, semiconducting thin-films comprising grains on the order of 5 μm in diameter. Thin-film X-ray diffraction indicates a monoclinic unit cell with molecules arranged in a herringbone pattern, which in conjunction with optical and atomic force microscopy indicate a thin-film with grains comprised of randomly oriented ZnTBP aggregates. Optical absorption measurements display broad absorption with bands characteristic of a D_4h symmetric porphyrin molecule. Organic field-effect transistors displayed field-effect mobilities on the order of 10^− 2 cm²/V s and ON-/OFF-current ratios exceeding 10².     

Tritium depth profile in matter using an imaging plate

A method to detect tritium non-destructively in regions deeper than the escape depth of beta rays is being developed using bremsstrahlung induced by beta rays with an imaging plate (IP). An IP made of europium-doped BaFBr(I), a photostimulated luminescence (PSL) material, is a two-dimensional radiation sensor. The bremsstrahlung energy spectrum is a continuum with photon energies, varying based on the atomic number and thickness of the target (or absorbing) material. When tritium migrates into matter, the bremsstrahlung energy spectrum distribution would change. The PSL intensity of the IP is affected by this energy spectrum variation. In order to quantify the amount of tritium in deeper regions with the IP technique, a tritium depth profile is required.In this study, a new method of obtaining a tritium depth profileusing the combined technique of the IP and thin absorbers is presented.     

Breeding of high malate‐producing diploid sake yeast with a homozygous mutation in the VID24 gene

Malate is an important taste component of sake (a Japanese alcoholic beverage) that is produced by the yeast Saccharomyces cerevisiae during alcoholic fermentation. A variety of methods for generating high malate‐producing yeast strains have been developed to date. We recently reported that a high malate‐producing strain was isolated as a mutant sensitive to dimethyl succinate (DMS), and that a mutation in the vacuolar import and degradation protein (VID) 24 gene was responsible for high malate productivity and DMS sensitivity. In this work, the relationships between heterozygous and homozygous mutants of VID24 and malate productivity in diploid sake yeast were examined and a method was developed for breeding a higher malate‐producing strain. First a diploid yeast was generated with a homozygous VID24 mutation by genetic engineering. The homozygous integrants produced more malate during sake brewing and grew more slowly in DMS medium than wild‐type and heterozygous integrants. Thus, the genotype of the VID24 mutation influenced the level of malate production and sensitivity to DMS in diploid yeast. Then a homozygous mutant from a heterozygous mutant was obtained without genetic engineering by ultraviolet irradiation and culturing in DMS with nystatin enrichment. The non‐genetically modified sake yeast with a homozygous VID24 mutation exhibited a higher level of malate productivity than the parent heterozygous mutant strain. These findings provide a basis for controlling malate production in yeast, and thereby regulating malate levels in sake. Copyright © 2016 The Institute of Brewing & Distilling