Apoptotic cells were quantified by flow cytometry, using an eBioscience Annexin V Apoptosis Detection Kit FITC (ThermoFisher Scientific). Jurkat cells were seeded in 12-well plates, then treated with PBS (vehicle) or 40 µg normal-derived exosomes and 40 µg RCC-derived exosomes for 24 h at 37 °C. Jurkat T cells were isolated and phosphate-buffered saline was then used to wash them twice. Cells were then stained, using the eBioscience Annexin V Apoptosis Detection Kit FITC (Invitrogen) outlined in the manufacturer’s instructions. All stained cells were processed by MACSQuant analyzer 10 and Flowlogic Analysis Software.
Ebioscience annexin 5 fitc apoptosis detection kit
Manufactured by Thermo Fisher Scientific
Sourced in United States
The EBioscience™ Annexin V-FITC Apoptosis Detection Kit is a tool used to detect and quantify apoptosis, a form of programmed cell death, in cell populations. The kit contains recombinant Annexin V conjugated to the fluorescent dye FITC, which binds to phosphatidylserine, a molecule that is exposed on the surface of apoptotic cells. The kit also includes a propidium iodide solution to distinguish between early apoptotic, late apoptotic, and necrotic cells.
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Lab products found in correlation
Facscalibur flow cytometer, by BD (4 mentions)
Attune nxt flow cytometer, by Thermo Fisher Scientific (3 mentions)
Facscanto 2, by BD (3 mentions)
Facscalibur, by BD (3 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
7 aad, by Thermo Fisher Scientific (2 mentions)
Facscan, by BD (2 mentions)
Inverted microscope, by Leica Microsystems (2 mentions)
Cellquest software, by BD (2 mentions)
Fxcycle pi rnase staining solution, by Thermo Fisher Scientific (2 mentions)
Propidium iodide solution, by Merck Group (2 mentions)
Potassium permanganate, by Merck Group (1 mentions)
Concentrated sulfuric acid, by Merck Group (1 mentions)
N isopropylacrylamide nipam, by Merck Group (1 mentions)
Rpmi medium, by Thermo Fisher Scientific (1 mentions)
Hydrochloric acid (hcl), by Merck Group (1 mentions)
Hydrogen peroxide, by Merck Group (1 mentions)
Sodium nitrate, by Merck Group (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
Streptomycin, by Thermo Fisher Scientific (1 mentions)
53 protocols using ebioscience annexin 5 fitc apoptosis detection kit
1
Exosome-Induced Apoptosis Quantification
2
Detecting Cell Apoptosis via Annexin V-FITC
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Cell apoptosis were detected at 48 h posttransfection using eBioscience™ Annexin V-FITC Apoptosis Detection Kit (Invitrogen) following manufacturer’s instruction. Generally, collected LoVo and HCT116 cells were resuspended in 200 μl Binding Buffer (1×), and then stained with 5 μl Annexin V-FITC and 10 μl Propidium Iodide (PI, 20μg/ml) for 10 min at room temperature in the dark. Next, apoptotic rates were determined using a flow cytometry (FACScan; BD Biosciences, San Jose, CA, USA).
3
Annexin V-FITC Apoptosis Assay
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Cellular apoptosis was assessed using the double staining of Annexin V-fluorescein isothiocyanate (Annexin V-FITC) and propidium iodide (PI) via the eBioscience™ Annexin V-FITC apoptosis detection kit (Invitrogen). Cell suspension (195 μL) in 1× Bing buffer (6×104 cells) was added with 5 μL of Annexin V-FITC for 10 min. PI (10 μL) was added to the tube after cell washing and resuspension with 190 μL of binding buffer, whereafter the apoptotic cells (Annexin V+/PI− and Annexin V+/PI+) were discerned by flow cytometry (BD Biosciences, San Diego, CA, USA). The apoptosis rate was accurately calculated using the general formula: apoptotic cells/total cells × 100%.
4
Annexin V-FITC Apoptosis Detection
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6 × 104 cells were collected for cell apoptosis detection through the double staining with Annexin V-fluorescein isothiocyanate (FITC) and propidium iodide (PI). The procedures were in accordance with the manufacturer’s specification of eBioscience™ Annexin V-FITC Apoptosis Detection Kit (Invitrogen) [25 (link)]. The apoptotic cells can be labeled by Annexin V (+)/PI (−) and Annexin V (+)/PI (+) on the flow cytometer (BD Biosciences, San Diego, CA, USA). The apoptotic rate was calculated using the formula: apoptotic cells/total cells × 100%.
5
Assessing Apoptosis in HepG2 Cells
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To assess the apoptotic rate, eBioscience™ Annexin V-FITC Apoptosis Detection kit (Invitrogen, Carlsbad, CA, USA) was used according to the manufacturer's guidelines. HepG2 cells were seeded in 6 well-plates and exposed to 40 μM PFOA and GenX (40, 100, 250, and 500 μM) for 12 h. The apoptotic rate of the sample after treatment was measured using a BD Accuri C6 Plus Flow Cytometer Analyzer (BD Biosciences, San Jose, CA, USA).
6
Evaluating Cell Apoptosis in RAW264.7 Cells
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For cell apoptosis evaluation, RAW264.7 cells were planted into 6-well plates with 2 × 105 cells each well for 24 h and then treated as Control, LPS (1 μg/ml), LPS (1 μg/ml) + TMP (5 or 10 μg/ml) and LPS (1 μg/ml) + Dex (5 μM) respectively. RAW264.7 cells were blown off with PBS from 6-well plates and washed in PBS two times. After incubation with PI and Annexin V with eBioscience™ Annexin V-FITC Apoptosis Detection Kit (Invitrogen, Carlsbad, CA, United States) according to instructions, the cells were analyzed using flow cytometry.
7
Graphene oxide-based nanoparticle synthesis
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Some reagents were used for the preparation of GO, including graphite, sodium nitrate (NaNO3), concentrated sulfuric acid (H2SO4, 95–98%), hydrochloric acid (HCl, 37%), potassium permanganate (KMNO4), and hydrogen peroxide (H2O2, 30%), purchased from Merck Co. (USA). For the polymerization reaction, two monomers, including N-isopropylacrylamide (NIPAM; 97%) and 1-vinyl-2-pyrrolidinone (VP; 95%) were obtained from Sigma-Aldrich Co. (USA). Two other regents, that is, 2,2′-azobis(2-methylpropionitrile) (AIBN) and lysine hydrochloride (Lys; 98%), were also provided by Merck Co. (USA).
All solvents were purified according to standard purification methods. FU, as an anticancer drug, was purchased from Sigma Aldrich Co. (USA). All materials used in the biological processes and assays, including 3-(4,5-dimethylthiazol-2-yl)-2,5-diphe-nyltetrazolium bromide (MTT), trypsin, fetal bovine serum (FBS), and RPMI medium, were purchased from Gibco BRL Life Technologies (USA). The eBioscience™ Annexin V-FITC Apoptosis Detection Kit was also purchased from Invitrogen Co. (USA). Finally, the A549 cells, as human lung cancer cells, were provided by the Cell Bank of Pasteur Institute of Iran (Tehran, Iran).
All solvents were purified according to standard purification methods. FU, as an anticancer drug, was purchased from Sigma Aldrich Co. (USA). All materials used in the biological processes and assays, including 3-(4,5-dimethylthiazol-2-yl)-2,5-diphe-nyltetrazolium bromide (MTT), trypsin, fetal bovine serum (FBS), and RPMI medium, were purchased from Gibco BRL Life Technologies (USA). The eBioscience™ Annexin V-FITC Apoptosis Detection Kit was also purchased from Invitrogen Co. (USA). Finally, the A549 cells, as human lung cancer cells, were provided by the Cell Bank of Pasteur Institute of Iran (Tehran, Iran).
8
Annexin V-FITC Apoptosis Assay
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Cell death was determined using the eBioscience™ Annexin V-FITC Apoptosis Detection Kit (Invitrogen, CA, USA), according to the manufacturer’s instructions. Briefly, cells were treated with SS as described above before harvesting, and resuspended in binding buffer with 5 μL Annexin V-FITC and propidium iodide (PI) (20 μg/mL) at room temperature for 10 min. The cells were centrifuged for 5 min at 2600× g and the pellets were resuspended in a binding buffer. Finally, the percentage of cell death was evaluated by flow cytometry (Beckman Coulter Inc., Brea, CA, USA). The cells positive for Annexin V-FITC and PI were counted as dead cells.
9
Neuroprotective Effects of Manganese on Dopaminergic Cells
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Dulbecco's modified Eagle's medium (DMEM), penicillin, streptomycin, and fetal bovine serum (FBS) were obtained from Gibco (Grand Island, NY, USA). Dopamine (DA), Manganese(II) chloride tetrahydrate (MnCl 2 •4H 2 O), MPP+ iodide (MPP + is active metabolite of MPTP), and LPS were purchased from Sigma-Aldrich (St. Louis, MO, USA). CellTiter 96 ® Aqueous One Solution Reagent was purchased from Promega (Promega, WI, USA), and the eBioscience™ Annexin V-FITC Apoptosis Detection Kit and Cell Cycle Detection Kit were obtained from Invitrogen (Thermo Fisher, Wa l t h a m , M A , U S A ) a n d K e y g e n ( K e y g e n Biotechnology, Jiangsu, China), respectively. The radioimmunoprecipitation assay (RIPA) reagent and BCA Protein Assay Kit were obtained from Beyotime Biotechnology (Beijing, China). The Easysee reagent was purchased from TransGen Biotech (Beijing, China). Antibodies against SCG3 (SC-50289), SCG2 (SC-50290), and β-actin (SC-47778) were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). RNAiso Plus, PrimeScript RT Reagent Kit, and SYBR Prime EX Taq were purchased from Takara Biotechnology (Shiga, Japan).
10
Evaluating Cell Death Mechanisms in Breast Cancer
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To determine the cell death mechanism of breast cancer cell lines and fibroblasts cells treated with ZnO. Annexin V/PI stain was examined by flow cytometry. MDA-MB-231/WT, MDA-MB-231/DR, MCF7/WT, MCF7/DR (2 × 105 cells/well) were seeded in 12-well plates (SPL, South Korea) and incubated at 37 °C for 24 h. Cells were then incubated in triplicate with 1 mL of compatible medium containing 10 µg/mL of ZnO for 24 h. Untreated cells were used as a negative control. Following treatment, cells were harvested using 250 µL of Accutase (Capricorn Scientific, Ebsdorfergrund, Germany) and Eagle’s Minimum Essential Medium (EMEM) (Euroclone SpA, Via Figino, Germany). According to the manufacturer’s instructions, the apoptosis assay was performed using eBioscience™ Annexin V-FITC Apoptosis Detection Kit (Invitrogen, Waltham, MA, USA). Samples were analyzed immediately using FACS Canto II (BD, San Jose, CA, USA).
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