Phen green sk

Manufactured by Thermo Fisher Scientific

Sourced in United States

Phen Green SK is a fluorescent dye that can be used to detect and quantify calcium levels in cells. It is a calcium-sensitive probe that emits fluorescence upon binding to calcium ions. The core function of Phen Green SK is to provide a reliable and sensitive method for measuring calcium concentrations in various biological samples.

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Lab products found in correlation

Accuri c6, by BD (2 mentions) Iron assay kit, by Merck Group (2 mentions) Lsm 780, by Zeiss (2 mentions) C11 bodipy581 591, by Thermo Fisher Scientific (1 mentions) H2dcfda, by BD (1 mentions) Bodipy 581 591 c11, by BD (1 mentions) H2dcfda, by Merck Group (1 mentions) Laser confocal microscope, by Leica (1 mentions) Microplate reader, by Agilent Technologies (1 mentions) Iron colorimetric assay kit, by Abcam (1 mentions) Fluorescence microscope, by Olympus (1 mentions) Eclipse te200 inverted microscope, by Nikon (1 mentions) Ixon charge coupled device camera, by Oxford Instruments (1 mentions) Chloroquine, by Merck Group (1 mentions) Bafilomycin, by Merck Group (1 mentions) D aminolevulinic acid, by Merck Group (1 mentions) Bodipy 581 591 c11, by Thermo Fisher Scientific (1 mentions) Gsk872, by Cayman Chemical (1 mentions) Necrosulfonamide, by Cayman Chemical (1 mentions) Ferrostatin 1, by Merck Group (1 mentions)

Spelling variants of this product

Phen Green SK diacetate (9 citations)

17 protocols using phen green sk

Quantification of Cellular Oxidative Stress

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Total ROS and lipid ROS were measured by flow analysis with H2DCFDA (Sigma) and C11‐BODIPY (581/591) (Invitrogen) staining according to the manufacturer's protocol, respectively. To analyze total ROS and lipid ROS, cells were stained with 25 × 10⁻⁶m H2DCFDA and 2 × 10⁻⁶m C11‐BODIPY(581/591) for 30 min at 37 °C, respectively, followed by using a flow cytometer (Accuri C6, BD Biosciences).
Measurement of cellular LIP using the fluorescent probe Phen Green SK (Invitrogen) has been described previously.^[⁴⁹^] Cells were incubated with 5 × 10⁻⁶m of Phen Green SK for 15 min in cell incubator. Then, cells were harvested and analyzed with a flow cytometer.

Guo J., Duan L., He X., Li S., Wu Y., Xiang G., Bao F., Yang L., Shi H., Gao M., Zheng L., Hu H, & Liu X. (2021). A Combined Model of Human iPSC‐Derived Liver Organoids and Hepatocytes Reveals Ferroptosis in DGUOK Mutant mtDNA Depletion Syndrome. Advanced Science, 8(10), 2004680.

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Inducing Iron Overload in Myocytes

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To induce iron overload, a membrane-permeable ferric ion (Fe³⁺)/8-hydroxyquinoline (8-HQ) complex (15 μM) was added to the myocyte bathing solution. The level of cytosolic iron was evaluated by the iron-sensitive dye Phen Green SK (Invitrogen by Thermo Fisher Scientific, Grand Island, NY, USA).

Siri-Angkul N., Song Z., Fefelova N., Gwathmey J.K., Chattipakorn S.C., Qu Z., Chattipakorn N, & Xie L.H. (2021). Activation of TRPC Channel Currents in Iron Overloaded Cardiac Myocytes. Circulation. Arrhythmia and electrophysiology, 14(2), e009291.

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Quantifying Cytosolic Iron Levels

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Cells were loaded with 20 μM Phen Green SK (Invitrogen) in culture medium at 37 °C for 20 min followed by a 15 min wash. Cellular fluorescence was excited at 488 nm and quenching of fluorescence was induced by the addition of the membrane-permeable transition metal chelator 2,2′-bipyridyl (5 mM). The 2,2′-bipyridyl analogue 4,4′-bipyridyl, which cannot bind or chelate Fe²⁺, was used as a control. Because many variables such as dye loading may contribute to the variation of basal fluorescence (F) of PG SK, the normalized change of fluorescence (ΔF/F) was used as readout to estimate the change in cytosolic Fe²⁺ levels.

Khan R.S., Baumann B., Dine K., Song Y., Dunaief J.L., Kim S.F, & Shindler K.S. (2019). Dexras1 Deletion and Iron Chelation Promote Neuroprotection in Experimental Optic Neuritis. Scientific Reports, 9, 11664.

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Quantifying Cellular Iron Levels

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According to the Iron Assay Kit (Sigma Aldrich, MAK025, USA), cells were added to iron assay buffer on ice and centrifuged at 16,000 g for 10 min at 4°C to obtain the supernatant. 50 μL of the supernatant was incubated with 50 μL of assay buffer in a 96 multi-well microplate for 30 min at 25°C. Samples were incubated with 100 μL of the iron probe for 60 min at 25°C while protected from light. The absorbance at 593 nm was measured using a microplate reader (BioTek, Winooski, VT). The final iron concentration was calculated by the corresponding formula. The chelatable intracellular iron pool was determined by using the fluorescent probe Phen Green SK (Invitrogen, P14313, USA). Cells were cultured on confocal dishes and loaded with 20 μM Phen Green SK for 30 min in PBS. The cells were then washed with PBS and imaged using a laser confocal microscope (Leica, Germany). The experiments were performed in triplicate.

Lan Y., Yang T., Yue Q., Wang Z., Zhong X., Luo X., Zuo B., Zhang M., Zeng T., Liu B, & Guo H. (2023). IRP1 mediated ferroptosis reverses temozolomide resistance in glioblastoma via affecting LCN2/FPN1 signaling axis depended on NFKB2. iScience, 26(8), 107377.

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Intracellular Iron Quantification Assays

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Intracellular Fe²⁺ levels were determined using an Iron Colorimetric Assay kit (BioVision, Inc., California, USA) and the fluorescent probe Phen green SK (Invitrogen, Carlsbad, CA, USA) strictly in line with the manufacturers’ instructions, respectively. For the colorimetric method, the absorbance at the wavelength of 593 nm was detected with a microplate reader. For the fluorescent probe method, the images were captured by a fluorescence microscope (Olympus, Tokyo, Japan).

Jin Y., Chen L., Li L., Huang G., Huang H, & Tang C. (2022). SNAI2 promotes the development of ovarian cancer through regulating ferroptosis. Bioengineered, 13(3), 6451-6463.

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Intracellular Iron Quantification in Myocytes

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To determine cytosolic ferrous iron (Fe²⁺) loading, ventricular myocytes were loaded with 40 µM Phen Green SK (Invitrogen) for 10 min at room temperature. In order to determine mitochondrial Fe²⁺, ventricular myocytes were loaded with 5 µM rhodamine B-[(1,10-phenanthroline-5-yl)-aminocarbonyl]benzyl ester (RPA; Squarix Biotechnology by Axxora, Recklinghausen, Germany) for 20 min at 37 °C. Fluorescence (Ex/Em: 484/520 nm for Phen Green and 543/560 nm for RPA) was monitored using an Eclipse TE200 inverted microscope (Nikon, Tokyo, Japan) and recorded using an Ixon Charge-Coupled Device (CCD) camera (Andor Technology, Concord, MA, USA).

Gordan R., Fefelova N., Gwathmey J.K, & Xie L.H. (2020). Iron Overload, Oxidative Stress and Calcium Mishandling in Cardiomyocytes: Role of the Mitochondrial Permeability Transition Pore. Antioxidants, 9(8), 758.

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Investigating Regulated Cell Death Pathways

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The following reagents were used: d-aminolevulinic acid, bafilomycin, chloroquine, ferrostatin-1, (Sigma), Laemmli buffer (GeneTex), Bodipy 581/591 C11, Phen Green SK, (Invitrogen), TMRE (Abcam), protoporphyrin IX (Frontier), GSK872, necrosulfonamide, (Cayman), Liproxstatin (Tocris), and Dynasore (Abcam). Antibodies used are listed in Supplementary Table 1 of the Supplementary Information File.

Lynch J., Wang Y., Li Y., Kavdia K., Fukuda Y., Ranjit S., Robinson C.G., Grace C.R., Xia Y., Peng J, & Schuetz J.D. (2023). A PPIX-binding probe facilitates discovery of PPIX-induced cell death modulation by peroxiredoxin. Communications Biology, 6, 673.

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Visualizing Labile Iron in Caco-2 Cells

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To visualize labile iron, microscopy imaging of the fluorescent dye Phen Green SK (Invitrogen, Eugene, OR, USA) was performed. Wild-type and DMT1 knockout Caco-2 cells were seeded at a density of 1 × 10⁴ cells/cm² in 6-well plates. After 14 days, the medium was discarded and the cells were washed twice with PBS and then incubated with DMEM containing 100 μM DFO. Twenty-four hours later, the medium was discarded and the cells were rinsed twice with PBS before being treated with DMEM containing different concentrations of Fe-Gly and FeSO₄ (0, 25, 50, 100, 200 μM). The cells were incubated for an additional 2 h, then rinsed twice with PBS and stained with PBS containing 10 μM Phen Green SK for 1 h at 37 °C. Next, the stained cells were rinsed with PBS and observed with a fluorescence microscope (IX7, Olympus Corporation, Tokyo, Japan).

Yu X., Chen L., Ding H., Zhao Y, & Feng J. (2019). Iron Transport from Ferrous Bisglycinate and Ferrous Sulfate in DMT1-Knockout Human Intestinal Caco-2 Cells. Nutrients, 11(3), 485.

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Quantifying Intracellular Iron Content

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Stable CHO cell lines expressing Egfp-Fpn Wt and Egfp-FpnK240E were grown to confluence on 6-cm dishes (PAA) using DMEM/GlutaMAX (Invitrogen) containing 800 mg/ml G418 (Invitrogen). Intracellular iron content was quantified by flow cytometry as previously reported [28 (link)]. Briefly, the cells were washed 3x with PBS and then incubated with 5 μM or 10 μM Phen Green SK (dipotassium salt; Invitrogen) diluted in PBS. After 20 min incubation, the cells were washed 3x with PBS and detached with PBS/0.5 mM EDTA. They were then centrifuged for 5 min at 2000 rpm and resuspended in 1 ml PBS for FACs analysis using a CyAn ADP flow cytometer (Beckman Coulter).

Bayele H.K, & Srai S.K. (2021). A disease-causing mutation K240E disrupts ferroportin trafficking by SUMO (ferroportin SUMOylation). Biochemistry and Biophysics Reports, 25, 100873.

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Quantifying Intracellular Chelatable Iron

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Intracellular chelatable iron was determined using the fluorescent indicator Phen Green SK from Thermo Fisher Scientific (P-14313). PSK-green was diluted with HBSS to 5 μM. Cells were washed twice with HBSS and incubated with 5 μM PSK-green at 37 degrees for 30 min. Then, the cells were washed with HBSS, digested with trypsin and collected by centrifugation. The fluorescence of Phen Green SK was detected using flow cytometry.

Yang J., Zhou Y., Xie S., Wang J., Li Z., Chen L., Mao M., Chen C., Huang A., Chen Y., Zhang X., Khan N.U., Wang L, & Zhou J. (2021). Metformin induces Ferroptosis by inhibiting UFMylation of SLC7A11 in breast cancer. Journal of Experimental & Clinical Cancer Research : CR, 40, 206.

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