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Kumamoto-shi, Japan

Nakazono M.,Kyushu University | Obayashi K.,Kumamoto University | Sasamoto K.,Dojindo Laboratories | Tomiyoshi K.,Kumamoto University | And 2 more authors.
Clinica Chimica Acta | Year: 2014

Background: Various styrylbenzene compounds were synthesized and evaluated as mainly Aβ amyloid sensors. These compounds, however, cannot be used for detecting amyloid deposition in peripheral nerves because of the inherent sensitivity of the compounds. These compounds often generate false positives especially in the basement membrane of blood vessels in histochemical studies. To overcome these problems, we must first synthesize other styryl compounds for detecting amyloid fibrils in tissues. Methods: A wide variety of symmetrical and unsymmetrical styrylbenzene derivatives were synthesized and then these compounds were used to detect amyloid fibrils in autopsy and biopsy samples from patients with various systemic and localized forms of amyloidosis such as familial amyloidotic polyneuropathy (FAP), senile systemic amyloidosis (SSA), amyloid A (AA) amyloidosis, localized AL amyloidosis, and Alzheimer's disease. Results: 1-Methoxy-2,5-bis-styrylbenzene and 2-(2-(2-fluoroethoxy)ethoxy)ethoxy)-2,5-bis-styrylbenzene (EEEFSB) detected amyloid fibrils in both in vitro and in vivo histopathological studies. 1-Methoxy-2,5-bis-styrylbenzene also showed a high strength of fluorescence with amyloid deposition in peripheral nerves in a patient with FAP. Conclusions: 1-Methoxy-2,5-bis-styrylbenzene and EEEFSB may prove a useful tool for diagnosing amyloidosis, not only in a histochemical study but also in whole body amyloid positron emission tomography (PET) imaging. © 2014 Elsevier B.V.

Uchida A.,University of Miyazaki | Sassa H.,Chiba University | Takenaka S.,University of Miyazaki | Takenaka S.,Dojindo Laboratories | And 3 more authors.
Plant OMICS | Year: 2012

The differences between stylar protein of the Japanese pear (Pyrus pyrifolia (Burm.f.)) cultivars 'Kosui' (S4S5) and 'Kikusui' (S2S4) were compared by two-dimensional difference gel electrophoresis (2-D DIGE), and were labelled and visualized with different fluorescent dyes (IC3-OSu, IC5-OSu) on a single 2-D gel. The individual different expressed proteins spots were subjected to identification. The proteins were analyzed by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF/MS) to identify proteins related to gametophytic self-incompatibility (GSI). S4-RNase and thaumatin-like protein 1 were successfully detected as expected in the pistils of 'Kosui' and 'Kikusui'. S5-RNase was also detected in the pistils of 'Kosui'. However, we could not detect S2-RNase in 'Kikusui' in this study, possibly because the level of expression of S2-RNase might be minuscule, or the estimated isoelectric point (pI) of S2-RNase (pI:9.26) was more basic than S4-RNase (pI: 9.17) and S5-RNase (pI: 9.01). These results indicate that proteomic studies are effective tools for detection of the expected proteins and might be helpful for finding the unknown key proteins related to the mechanism of self-incompatibility (SI) in many other SI plants.

Uchida A.,University of Miyazaki | Takenaka S.,University of Miyazaki | Takenaka S.,Dojindo Laboratories | Sakakibara Y.,University of Miyazaki | And 5 more authors.
Journal of the Japanese Society for Horticultural Science | Year: 2012

In Citrus, self-incompatibility (SI) regulation is gametophytic, and this phenomenon is an economically critical problem for some Citrus cultivars without high parthenocarpic ability. Few molecular biological studies of SI in Citrus have been performed, and the molecular mechanism of SI has not been clarified. To investigate the effect of different stages of style development on pollen tube behavior, flower buds of self-incompatible 'Hyuganatsu' (Citrus tamurana hort. ex Tanaka) were histologically assayed. When the flower buds were observed 168 hours after pollination, pollen tubes in the self-pollinated flower buds 1 and 3 days before anthesis (DBA) were arrested in the upper part of the styles, while those in flower buds self-pollinated 5 DBA reached the base of the styles. These results revealed that SI in 'Hyuganatsu' has not yet occurred in the flower buds 5 DBA, but generated in the flower buds 3 DBA. To search SI-related pistil proteins in Citrus, we profiled a number of protein expressions in 'Hyuganatsu' styles of 1, 3, and 5 DBA by two-dimensional gel electrophoresis and MALDI-TOF mass spectrometry. One-hundred thirty-eight protein spots were significantly different in abundance among the three stages, and 17 up-regulated and 26 down-regulated protein spots could be identified. Among the 17 up-regulated proteins, nine up-regulated proteins exhibited the expression pattern of 1 DBA ≥ 3 DBA > 5 DBA, a pattern which reflected the transmission from SC to SI, evaluated by pollen tube growth. BLASTP homology search against the peptide sequences of these proteins was carried out and predicted the proteins related to the maintenance of cell shape, the signaling of pollen tube growth, the flavonol biosynthesis, the responses to various stresses, photosynthesis, and the methionine metabolic process. Among the nine proteins, some may be the SI-related pistil proteins; however, it was not possible to determine the SI-related key protein of the style in Citrus. © 2012.

Tsukatani T.,Fukuoka Industrial Technology Center | Suenaga H.,Fukuoka Industrial Technology Center | Shiga M.,Dojindo Laboratories | Matsumoto K.,Sojo University
Letters in Applied Microbiology | Year: 2014

A rapid microplate method for the proliferation assay of fungi and the antifungal susceptibility testing using the colorimetric microbial viability assay based on the reduction in a tetrazolium salt 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium, monosodium salt (WST-8) with 2-methyl-1,4-napthoquinone as the electron mediator was developed. The proposed method was useful to measure the proliferation of 18 kinds of moulds and seven kinds of yeasts, including representative pathogens such as Aspergillus spp., Candida spp. and Cryptococcus spp. Linear relationships between the absorbance and viable fungal cell density were obtained for all fungi, suggesting that the absorbance change reflected the fungal proliferation. In addition, the minimum inhibitory concentrations (MICs) against a variety of different pathogenic moulds and yeasts for amphotericin B, itraconazole and 5-flucytosine were determined by susceptibility testing using the proposed method and compared with those obtained using the conventional broth microdilution method. There was an excellent agreement between the results obtained using the WST-8 colorimetric method and those obtained using the conventional Clinical and Laboratory Standard Institute method. The WST-8 colorimetric assay is a useful method for rapid determination of accurate MICs for a variety of different fungi. © 2014 The Society for Applied Microbiology.

Tsukatani T.,Fukuoka Industrial Technology Center | Suenaga H.,Fukuoka Industrial Technology Center | Ishiyama M.,Dojindo Laboratories | Ezoe T.,Dojindo Laboratories | Matsumoto K.,Kyushu University
Food Chemistry | Year: 2011

A method for the determination of water-soluble vitamins using a colorimetric microbial viability assay based on the reduction of the tetrazolium salt {2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H- tetrazolium, monosodium salt (WST-8)} via 2-methyl-1,4-napthoquinone (NQ) was developed. Measurement conditions were optimized for the microbiological determination of water-soluble vitamins, such as vitamin B 6, biotin, folic acid, niacin, and pantothenic acid, using microorganisms that have a water-soluble vitamin requirement. A linear relationship between absorbance and water-soluble vitamin concentration was obtained. The proposed method was applied to determine the concentration of vitamin B 6 in various foodstuffs. There was good agreement between vitamin B 6 concentrations determined after 24 h using the WST-8 colorimetric method and those obtained after 48 h using a conventional method. The results suggest that the WST-8 colorimetric assay is a useful method for the rapid determination of water-soluble vitamins in a 96-well microtiter plate. © 2011 Elsevier Ltd. All rights reserved.

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