ARPE-19 cells (CRL-2302) were purchased from the American Type Culture Collection (Manassas, VA, USA) and routinely cultured in Dulbecco’s Modified Eagle Medium/F-12 supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin (WELGENE Inc., Gyeongsan, Republic of Korea) as described previously [27 (link)]. To investigate beneficial effects of phloroglucinol on oxidative damage, cells were cultured in media containing desired concentrations of phloroglucinol and H2O2 (Thermo Fisher Scientific, Inc., Waltham, MA, USA) for 24 h or pretreated with phloroglucinol, N-acetyl-L-cysteine (NAC), Mito-TEMPO, and/or 3-methyladenine (3-MA, Sigma-Aldrich Co., St. Louis, MO, USA) for 1 h prior to treatment with H2O2 for 24 h. To investigate the blocking effect of phloroglucinol on the generation of ROS induced by H2O2, cells were pretreated with phloroglucinol, NAC, and Mito-TEMPO for 1 h and then treated with H2O2 for 1 h. To acquire fluorescence images of ROS generation, γH2AX expression, and autophagic vacuoles, cells cultured on coverslips were stimulated with H2O2 in the presence or absence of phloroglucinol, NAC, and/or Mito-TEMPO. After treatment, cells were washed with phosphate-buffered saline and subjected to fluorescence staining.
Phloroglucinol
Manufactured by Thermo Fisher Scientific
Sourced in United States
Phloroglucinol is a chemical compound commonly used as a reagent in analytical chemistry. It is a white crystalline solid that is soluble in water and organic solvents. Phloroglucinol is primarily used as a staining agent in histology and microscopy applications to detect the presence of certain compounds or structures in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
3 methyladenine 3 ma, by Merck Group (1 mentions)
Penicillin streptomycin, by Welgene (1 mentions)
Phloroglucinol, by Merck Group (1 mentions)
N acetyl l cysteine, by Merck Group (1 mentions)
Mitotempo, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Welgene (1 mentions)
Arpe 19 cells crl 2302, by American Type Culture Collection (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)
Methyl thiazolyl tetrazolium (mtt), by Thermo Fisher Scientific (1 mentions)
Isopropyl alcohol, by Merck Group (1 mentions)
Proteinase k, by Merck Group (1 mentions)
Rnase a, by Merck Group (1 mentions)
Caspase 3 activity assay kit, by Merck Group (1 mentions)
Mitochondrial fractionation kit, by Thermo Fisher Scientific (1 mentions)
Immobilon p pvdf membrane, by Merck Group (1 mentions)
Annexin 5 fitc apoptosis detection kit, by Abcam (1 mentions)
Primary and horseradish peroxidase conjugated secondary antibodies, by Cell Signaling Technology (1 mentions)
Hydrochloric acid (hcl), by Merck Group (1 mentions)
Ammonium fluoride nh4f, by Thermo Fisher Scientific (1 mentions)
Milli q, by Merck Group (1 mentions)
7 protocols using phloroglucinol
1
Phloroglucinol Protects ARPE-19 Cells from Oxidative Stress
2
Antimethanogenic Chloroform-Cyclodextrin Matrix
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The antimethanogen compound used was a halogenated hydrocarbon (chloroform) entrapped in a β-cyclodextrin matrix (6–7% w/w chloroform) as described by Martinez-Fernandez et al. (2016) (link). The phenolic compound used was phloroglucinol (Thermo Fisher Scientific, Scoresby, VIC, Australia; purity of 99%).
3
Phloroglucinol-Mediated Cytoprotective Mechanisms
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All materials necessary for cell culture were obtained from WELGENE Inc. (Gyeongsan, Republic of Korea). Phloroglucinol (1,3,5-trihydroxybenzene), H2O2, MTT, JC-1, mitochondrial fractionation kit, and enhanced chemiluminescence (ECL) solution were purchased from Thermo Fisher Scientific (Waltham, MA, USA). Dimethyl sulfoxide (DMSO), ZnPP, DCF-DA, DAPI, RNase A, proteinase K, isopropyl alcohol, EtBr, and caspase-3 activity assay kit were obtained from Sigma-Aldrich Co. (St. Louis, MO, USA). The comet assay kit was purchased from Trevigen, Inc. (Gaithersburg, MD, USA). HO-1 enzyme-linked immunosorbent assay (ELISA) kit and annexin V-FITC apoptosis detection kit were purchased from Abcam, Inc. (Cambridge, UK). Immobilon®-P PVDF membranes were obtained from Merck Millipore (Bedford, MA, USA). Primary and horseradish peroxidase-conjugated secondary antibodies were purchased from Cell Signaling Technology (Beverly, MA, USA), Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA), and Abcam, Inc (Cambridge, UK).
4
Phlorotannin Content Determination
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The total phlorotannin content was determined according to the method of Singleton and Rossi (1965) . The standard was 98% phloroglucinol (PGE, 50–300 μg/mL) from Thermo Fisher Scientific (Waltham, MA, USA). A mixture of 200 μL of subfraction sample, 2.6 mL distilled water, and 200 μL of 2 M Folin-Ciocalteu reagent were incubated at room temperature for 6 min. The reaction was stopped by adding 2 mL of 7% (w/v) Na2CO3. The mixture was incubated again at room temperature for 90 min, and the absorbance at 750 nm was measured with a UV-VIS Spectrophotometer (V-530; Jasco, Tokyo, Japan). Total phlorotannin content was calculated from the regression equation of the standard curve from phloroglucinol.
5
Fabrication of Alkaline Fuel Cell Electrodes
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Phloroglucinol, ammonium fluoride (NH4F) and Pluronic F-127 were procured from Acros Organics, India. Gas diffusion layers (GDL, Sigracet DC-35, Germany) were used as electrodes. The Pt/C catalyst (40 wt% Pt supported on Vulcan carbon) was obtained from Johnson-Matthey, UK and was used as a catalyst layer over the GDL surface. Ethanol, formaldehyde (37–41%) and hydrochloric acid (HCl) were purchased from Merck, Germany. Nafion (5 wt%) ionomer was obtained from Du Pont, USA. Fumion-FAA-3 (10 wt%) ionomer solution (polyaromatic backbone with terminal quaternary ammonium ions and bromide counter ions) was procured from Fumatech, Germany. All the chemicals employed in this work were analytical grade and used without any further decontamination process. Milli-Q water (resistivity is 18.2 mΩ cm) was utilized throughout the experiments.
6
Antioxidant Evaluation of Chitosan-Based Biomaterials
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Crude FFv, CS (low molecular weight, 75–85% deacetylation degree), gelatin from porcine skin (type A), barium chloride, trichloroacetic acid, 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox), glacial acetic acid, Dulbecco’s Modified Eagle’s (DMEM), L-glutamine, non-essential amino acids, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) were purchased from Sigma-Aldrich (Burghausen, Germany). Potassium sulphate (K2SO4) and Folin-Ciocalteau reagent were from Panreac AppliChem (Barcelona, Spain). 2,2-azinobis(3-ethyl-benzothiazoline-6-sulfonate) (ABTS) was supplied by Alfa Aesar (Kandel, Germany), and sodium carbonate (Na2CO3) was purchased from Merck (Darmstadt, Germany). Phloroglucinol was supplied by AcrosOrganics (Shanghai, China). Polystyresulfonate sodium salt (PSS) standards were acquired from Sigma and Fluka (Barcelona, Spain and St. Louis, MO, USA, respectively). Phosphate-buffered saline (PBS) tablets pH 7.4 were acquired from VWR (Rosny-sous-Bois, France) and dimethyl sulfoxide (DMSO) from Carlo Erba (Sabadell, Spain). Penicillin/streptomycin and fetal bovine serum (FBS) were provided by Gibco (Life Technologies, Carlsbad, CA, USA), while the lactate dehydrogenase (LDH) kit was purchased from Takara Bio (Tokyo, Japan). Ultrapure water (Milli-Q, Millipore, Watford, UK) was used throughout. All other chemicals were reagent grade.
7
Lignin and Callose Response to ES20
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To examine the effect of ES20 on lignin and callose accumulation, Col-0 seedlings were grown on 1/2 MS media supplemented with DMSO (0.1%) or ES20 (4 M) in dark condition for three days. Lignin staining was performed following published protocol (Pradhan Mitra and Loque, 2014). Dark-grown seedlings were incubated in phloroglucinol (Acros Organic) solution (20 mg/mL in ethanol: hydrocholoric acid (2:1 vol/vol)) for 5 min and then imaged under white light. Callose staining was performed by following published protocol (Harris et al., 2012) . Dark-grown seedlings were incubated in aniline blue (Acros Organic) staining solution (0.1 mg/mL in 0.07 M sodium phosphate buffer, pH 9) in the dark for 20 min. The seedlings were then imaged under UV light.
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