Samples of adipose tissue, pancreas, and liver were fixed in 4% paraformaldehyde in phosphate buffer solution (Wako) for 24 h, embedded in paraffin, sectioned at 4 μm, and stained with hematoxylin and eosin. For immunohistochemistry, paraffin-embedded sections were stained with monoclonal anti-UCP1 (dilution, 1:200; Abcam, Cambridge, MA, USA), anti-insulin (Histofine, Nichirei, Tokyo, Japan), anti-Ki67 (dilution, 1:1000; Abcam, Cambridge, MA, USA), and anti-F4/80 (AbD Serotec, Oxford, UK) antibodies. Images were acquired by using an FSX100 system (Olympus, Tokyo, Japan), and the UCP1 and F4/80 areas were evaluated by using cellSens Dimension software version 1.6 (Olympus). Histologic images were analyzed to calculate the sizes of adipocytes and insulin- and Ki67-positive areas by using Image J software (National Institutes of Health, Bethesda, MD, USA). For the analysis, 5 random images were captured for every sample.
Phosphate buffer solution (pbs)
Manufactured by Fujifilm
Sourced in Japan
Phosphate buffer solution is a laboratory reagent used to maintain a stable pH environment. It is a mixture of sodium phosphate and potassium phosphate salts dissolved in water. The solution helps to control the acidity or basicity of a system, ensuring consistent pH levels for various experimental and analytical applications.
Automatically generated - may contain errors
Lab products found in correlation
Fsx100 system, by Olympus (2 mentions)
Anti ki67, by Abcam (2 mentions)
Anti f4 80, by Bio-Rad (2 mentions)
Bz x710, by Keyence (2 mentions)
Anti ucp1, by Abcam (1 mentions)
Cellsens dimension software version 1, by Olympus (1 mentions)
F4 80, by Olympus (1 mentions)
Dylight 488, by Thermo Fisher Scientific (1 mentions)
Hoechst 33258, by Thermo Fisher Scientific (1 mentions)
Paraformaldehyde (pfa), by Fujifilm (1 mentions)
Mayer s hematoxylin, by Fujifilm (1 mentions)
Eosin y, by Fujifilm (1 mentions)
Histofine, by Nichirei Biosciences (1 mentions)
Cellsens dimension software, by Olympus (1 mentions)
Glutaraldehyde, by Electron Microscopy Sciences (1 mentions)
T butyl alcohol, by Fujifilm (1 mentions)
Auto fine coater, by JEOL (1 mentions)
Jsm 6510lv, by JEOL (1 mentions)
7 protocols using phosphate buffer solution (pbs)
1
Histological Analysis of Adipose, Pancreas, and Liver
2
Measuring Hydrogen's Effect on Phosphate Buffer
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The effect of H2 at low partial pressure was evaluated using the ORP of a phosphate buffer solution (10 mM at pH = 7.1; Fujifilm Wako Pure Chemical Corp., Osaka, Japan). For a mixture of gases at low pressure, the activity is equal to the ratio of the partial pressure of the gas over the standard pressure. The standard H2 gas contains 100 ppm of H2 ( in medical air, and the air contains 0.6 ppm of H2 ( ). The phosphate-buffered solution was not aerated before the experiment, although tap water has a positive ORP ranging from + 200 to + 400 mV due to dissolved O2. The air was bubbled in a phosphate-buffered solution at a rate of 0.5 L/min using a humidifier water bottle for 60 min at 22 °C. The air was supplied from the integrated piping system at the TMG Asaka Medical Center (Air Water Inc., Osaka, Japan), and standard H2 gas for calibration (100 ppm of H2 in medical air; Air Water Inc., Osaka, Japan) was supplied from the cylinder with a pressure regulator. The ORP and pH were measured before and after the air bubbling using an ORP/pH meter (pH6600 and ORP6600S; Custom, Tokyo, Japan).
3
Immunohistochemical Analysis of Gastric Tissues
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Gastric tissues were fixed using 4% paraformaldehyde in phosphate buffer solution (Wako Pure Chemical Industries) at 4 °C overnight. Next, these tissues were immersed in 15% sucrose in PBS at 4 °C overnight, frozen in dry ice powder, and sliced into 10-μm-thick sections. Sections were incubated with antibodies against HKα (1:100; rabbit polyclonal; Calbiochem, La Jolla, CA, USA) and HKβ (1:100; mouse monoclonal; Sigma-Aldrich), and then subsequently incubated with DyLight® 488 and DyLight® 649-labelled secondary antibodies (Invitrogen, Carlsbad, CA, USA). Hoechst 33258 (Invitrogen) was used for nuclear staining. The immunostained specimens were observed with a fluorescence microscope (BZ-X710; Keyence, Osaka, Japan).
4
Histological Analysis of Gastric Tissues
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Gastric tissues were fixed using 4% paraformaldehyde in phosphate buffer solution (Wako Pure Chemical Industries) at 4 °C overnight. Next, these tissues were immersed in 15% sucrose in PBS at 4 °C overnight, frozen in dry ice powder, and sliced into 10-μm-thick sections. Sections got fully dipped with Mayer’s hematoxylin (Wako Pure Chemical Industries) and eosin Y (Wako Pure Chemical industries), sequentially. The HE stained specimens were observed with a light field microscopy (BZ-X710; Keyence).
5
Histological Analysis of Metabolic Tissues
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Samples of adipose tissue, pancreas, and liver were fixed in 4% paraformaldehyde in phosphate buffer solution (Wako) for 24 h at room temperature, embedded in paraffin, sectioned at 4 μm, and stained with haematoxylin and eosin. For immunohistochemistry, paraffin-embedded sections were stained with monoclonal anti-insulin (Histofine; Nichirei, Tokyo, Japan), anti-Ki67 (dilution, 1:1000; Abcam, Cambridge, MA, USA), and anti-F4/80 (AbD Serotec, Oxford, UK) antibodies. Images were acquired by using an FSX100 system (Olympus, Tokyo, Japan), and the F4/80-positive area was quantified by using cellSens Dimension software (version 1.6, Olympus). Histologic images were analysed to calculate the sizes of adipocytes and of the insulin- and Ki67-positive areas by using Image J software (National Institutes of Health, Bethesda, MD, USA). For the analyses, 5 random images were captured from each sample.
6
Formulation and Characterization of NFT-Loaded Polymeric Micelles
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NFT was purchased from Tokyo Chemical Industry (Tokyo, Japan). Phosphate buffer solution was purchased from FUJIFILM Wako Pure Chemical (Osaka, Japan) and diluted with pure water to obtain a final concentration and pH of 25 mM and pH 6.8, respectively. Vinylpyrrolidone-vinyl acetate copolymer (Kollidon VA64, PVPVA), Eudragit (poly(methacrylic acid-co-methyl methacrylate)) L100-55 (Eudragit), and hydroxypropyl methylcellulose acetate succinate (MG grade, HPMCAS) were supplied by BASF (Ludwigshafen am Rhein, Germany), Evonik (Essen, Germany), and Shin-Etsu Chemical (Tokyo, Japan), respectively. Blank simulated intestinal fluid (SIF) was prepared by dissolving 10 mM sodium chloride and 28.4 mM monobasic sodium phosphate in pure water, which was adjusted to pH 6.5 via the addition of sodium hydroxide solution, where FeSSIF or FaSSIF powder for humans (Biorelevant, London, UK) was dissolved to prepare 3 or 15 mM SIF. All other chemicals were of reagent grade.
7
SEM Analysis of Root Canal Dentin-Resin Interface
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Additional samples of the Direct/DC and the Direct/PV groups were prepared from six caries-free human teeth (n = 3 in each group). The samples were divided into two parts parallel to the tooth axis, and the cut surfaces were treated with phosphoric acid for 15 s, washed with water, dried, and subsequently treated with sodium hypochlorite for 60 s, washed with water, and dried. After being fixed with a solution prepared by mixing 8% glutaraldehyde (Electron Microscopy Sciences, Philadelphia, PA, USA) and 0.2 mol/l cacodylate buffer solution (Fujifilm Wako Pure Chemical Corporation, Osaka, Japan) at 1:1, for more than 2 hours, the samples were washed several times with phosphate buffer solution (Fujifilm Wako Pure Chemical Corporation) and fixed with osmic acid solution (Nisshin EM Co., Ltd., Tokyo, Japan) for 2 hours. The samples were then washed again with phosphate buffer solution. Dehydration was subsequently carried out using an ethanol ascending system, and substitution was carried out with t-butyl alcohol (Fujifilm Wako Pure Chemical Corporation), followed by lyophilization. The dried samples were gold-evaporated (Auto Fine Coater, JEOL Ltd., Tokyo, Japan). The prepared samples were observed at the root canal dentin-resin interface using a scanning electron microscope (SEM, JSM-6510LV, JEOL Ltd.).
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