Kumamoto-shi, Japan
Kumamoto-shi, Japan

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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.


Tsukatani T.,Fukuoka Industrial Technology Center | Suenaga H.,Fukuoka Industrial Technology Center | Shiga M.,Dojindo Laboratories | Noguchi K.,Dojindo Laboratories | And 3 more authors.
Journal of Microbiological Methods | Year: 2012

The minimum inhibitory concentrations (MICs) obtained from the susceptibility testing of various bacteria to antibiotics were determined by a colorimetric microbial viability assay based on reduction of a tetrazolium salt {2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2. H-tetrazolium, monosodium salt (WST-8)} via 2-methyl-1,4-napthoquinone as an electron mediator and compared with those obtained by the broth microdilution methods approved by the Clinical and Laboratory Standard Institute (CLSI). Especially for drug-resistant bacteria, the CLSI method at an incubation time of 24. h tended to give lower MICs. The extension of incubation time was necessary to obtain consistent MICs for drug-resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococi (VRE) and multi-drug resistant Pseudomonas aeruginosa (MDRP) in the broth microdilution method. There was excellent agreement between the MICs determined after 24. h using the WST-8 colorimetric method and those obtained after 48-96. h using the broth microdilution method. The results suggest that the WST-8 colorimetric assay is a useful method for rapid determination of consistent MICs for drug-resistant bacteria. © 2012 Elsevier B.V..


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.


Sano K.,U.S. National Cancer Institute | Nakajima T.,U.S. National Cancer Institute | Miyazaki K.,Dojindo Laboratories | Ohuchi Y.,Dojindo Laboratories | And 3 more authors.
Bioconjugate Chemistry | Year: 2013

The ability to switch optical imaging probes from the quenched (off) to the active state (on) has greatly improved target to background ratios. The optimal activation efficiency of an optical probe depends on complete quenching before activation and complete dequenching after activation. For instance, monoclonal antibody-indocyanine green (mAb-ICG) conjugates, which are promising agents for clinical translation, are normally quenched, but can be activated when bound to a cell surface receptor and internalized. However, the small fraction of commonly used ICG derivative (ICG-Sulfo-OSu) can bind noncovalently to its mAb and is, thus, gradually released from the mAb leading to relatively high background signal especially in the liver and the abdomen. In this study, we re-engineered a mAb-ICG conjugate, (Panitumumab-ICG) using bifunctional ICG derivatives (ICG-PEG4-Sulfo-OSu and ICG-PEG8-Sulfo-OSu) with short polyethylene glycol (PEG) linkers. Higher covalent binding (70-86%) was observed using the bifunctional ICG with short PEG linkers resulting in less in vivo noncovalent dissociation. Panitumumab-ICG conjugates with short PEG linkers were able to detect human epidermal growth factor receptor 1 (EGFR)-positive tumors with high tumor-to-background ratios (15.8 and 6.9 for EGFR positive tumor-to-negative tumor and tumor-to-liver ratios, respectively, at 3 d postinjection). © 2013 American Chemical Society.


PubMed | Dojindo Laboratories, Kumamoto University, Curtin University Australia, Tokushima University and 2 more.
Type: Journal Article | Journal: Biochemical and biophysical research communications | Year: 2016

Recently, hydropersulfide (RSSH) was found to exist in mammalian tissues and fluids. Cysteine hydropersulfide can be found in free cysteine residues as well as in proteins, and it has potent antioxidative activity. Human serum albumin (HSA) is the most abundant protein in mammalian serum. HSA possesses a free thiol group in Cys-34 that could be a site for hydropersulfide formation. HSA hydropersulfide of high purity as a positive control was prepared by treatment of HSA with Na


PubMed | Kumamoto University, Japan Science and Technology Agency, Dojindo Laboratories, Showa Pharmaceutical University and Tohoku University
Type: | Journal: Nucleic acids research | Year: 2016

The 2-methylthio (ms


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.


Yamanaka K.,Doshisha University | Saito Y.,Doshisha University | Sakiyama J.,Dojindo Laboratories | Ohuchi Y.,Dojindo Laboratories | And 2 more authors.
RSC Advances | Year: 2012

Spy-LHP was developed as a fluorescent probe for the detection and imaging of lipid hydroperoxides in living cells. Although Spy-LHP detection of lipid hydroperoxides is sensitive and selective, this probe is unsuitable for live-cell imaging because of its high hydrophobicity. To overcome this limitation, 2-(4-diphenylphosphanyl-phenyl)-9-(3,6,9,12-tetraoxatridecyl)- anthra[2,1,9-def:6,5,10-d′e′f′]diisoquinoline-1,3,8, 10-tetraone, Liperfluo, was developed. Liperfluo is structurally similar to Spy-LHP, but is much more soluble in various organic solvents, such as ethanol and dimethylsulfoxide. The probe was successfully used to image lipid hydroperoxides in SH-SY5Y cells in which lipid peroxidation had been stimulated with 2,2′-azobis[2-(2-imidazolin-2-yl)propane] dihydrochloride or cumene hydroperoxide by using fluorescent microscopy and confocal laser scanning microscopy. Additionally, flow cytometric analysis showed that Liperfluo was four times more sensitive in detecting lipid hydroperoxides than Spy-LHP. These results suggest that Liperfluo is a useful fluorescent probe for investigating the roles of lipid peroxidation in a variety of cell pathophysiologies. This journal is © 2012 The Royal Society of Chemistry.


Hara S.,Tokyo Institute of Technology | Tatenaka Y.,Dojindo Laboratories | Ohuchi Y.,Dojindo Laboratories | Hisabori T.,Tokyo Institute of Technology | Hisabori T.,Japan Science and Technology Agency
Biochemical and Biophysical Research Communications | Year: 2015

The redox states of proteins in cells are key factors in many cellular processes. To determine the redox status of cysteinyl thiol groups in proteins in vivo, we developed a new maleimide reagent, a photocleavable maleimide-conjugated single stranded DNA (DNA-PCMal). The DNA moiety of DNA-PCMal is easily removed by UV-irradiation, allowing DNA-PCMal to be used in Western blotting applications. Thereby the state of thiol groups in intracellular proteins can be directly evaluated. This new maleimide compound can provide information concerning redox proteins in vivo, which is important for our understanding of redox networks in the cell. © 2014 Elsevier Inc. All rights reserved.


PubMed | Dojindo Laboratories
Type: Journal Article | Journal: Cytotechnology | Year: 2012

Human-human hybridoma cells secreting a human monoclonal antibody were cultured in a serum-free medium containing various organic pH buffers in order to clarify their effects on cell growth and antibody production. Organic pH buffers having either one sulfonic acid and several acyclic amine moieties, or several cyclic amine moieties containing two amino nitrogen did not inhibit cell growth; while other organic buffers sulfonic acid moiety plus several cyclic amine moieties containing one amino nitrogen slightly decreased cell growth, but enhanced antibody production. Using Fujitas organic conceptual diagram, a relationship between the organicity and inorganicity of a pH buffer to cell growth and antibody production was found. pH buffers with large inorganicity and small organicity values were favorable for cell growth, and buffers with small inorganicity and large organicity values were preferred to enhance antibody production. Although the pH buffering range affects cell growth, its effect on antibody production is not clear. In conclusion, 2-morpholinoethanesulfonic acid (MES), 3-morpholino-propanesulfonic acid (MOPS) and 1, 2-N, N-bis[N, N-di(2-sulfonoethyl)piperazinyl]ethane (Bis-PIPES) are shown to be the most optimal of the buffers tested, because they enhanced antibody production without decreasing the cell growth among the pH buffers tested here.

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