Cytotoxicity of PTT was determined by quantification of apoptotic or necrotic cancer cells. MDA-MB-231 cells were seeded at a density of 4 × 105 cells/well in 24-well plates for 24 h at 37 °C and they were then treated with or without CS-PPy NCs (210 μg/mL) for 4 h. After washing, the cells were treated with or without the 808-nm laser at 2 W/cm2 for 5 min. Subsequently, cells were cultured for additional 2 h and they were then harvested. Fluorescein isothiocyanate Annexin V Apoptosis Detection Kit (BD Biosciences, USA) was used to detect and quantify apoptosis using a flow cytometer (BD FACSVerse, NJ, USA). Annexin V-positive and PI-negative cells were recorded as apoptotic. Double-stained cells were considered to be apoptotic or necrotic.
Annexin 5 fluorescein isothiocyanate apoptosis detection kit
Manufactured by BD
Sourced in United States
The Annexin V-fluorescein isothiocyanate apoptosis detection kit is a laboratory product that enables the detection and quantification of apoptosis, a type of programmed cell death, in cell samples. The kit utilizes Annexin V, a protein that binds to phosphatidylserine, a molecule that is exposed on the surface of cells undergoing apoptosis. Annexin V is conjugated to the fluorescent dye fluorescein isothiocyanate (FITC), allowing for the visualization and analysis of apoptotic cells using flow cytometry or fluorescence microscopy.
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Lab products found in correlation
Facscalibur, by BD (5 mentions)
Facscalibur flow cytometer, by BD (5 mentions)
Accuri c6 flow cytometer, by BD (3 mentions)
Propidium iodide, by BD (3 mentions)
Accuri c6, by BD (2 mentions)
Cellquest pro software, by BD (2 mentions)
Cellquest software, by BD (2 mentions)
Facscan, by BD (2 mentions)
Facsverse, by BD (2 mentions)
Facsan flow cytometry, by BD (2 mentions)
Moflo xdp, by Beckman Coulter (2 mentions)
Epics xl mcl, by Beckman Coulter (2 mentions)
Dm2500, by Leica (1 mentions)
Accuri c5 flow cytometer, by BD (1 mentions)
Cell culture insert, by Corning (1 mentions)
Flow cytometry, by Thermo Fisher Scientific (1 mentions)
Annexin 5, by BD (1 mentions)
Spectra190, by Molecular Devices (1 mentions)
Propidium iodide, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Atlanta Biologicals (1 mentions)
53 protocols using annexin 5 fluorescein isothiocyanate apoptosis detection kit
1
Cytotoxicity Assay of Photothermal Cancer Treatment
2
Annexin V Apoptosis Assay by Flow
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To detect the translocation of phosphatidylserine, one of the markers of apoptosis, from the inner to the outer leaflet of the plasma membrane, cells were stained with Annexin V according to the manufacturer’s protocol of the fluorescein isothiocyanate Annexin V Apoptosis Detection Kit (BD Biosciences, San Diego, CA). Flow cytometry was performed using an Accuri C6 flow cytometer (BD Biosciences).
3
Quantifying Podocyte Apoptosis by Flow Cytometry
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The ratio of apoptotic cells was determined using a fluorescein isothiocyanate-Annexin V Apoptosis Detection kit (BD Biosciences). The analysis was performed as previously described (9 (link)). Briefly, the cells were stained with FITC-Annexin V and propidium iodide, according to the manufacturer's instructions. Podocyte apoptosis was determined by flow cytometry (FACSCalibur; BD Biosciences) and analyzed using Accuri C6 software (version 1.0.264.21; BD Biosciences). The presence of apoptotic cells in the tissue sections was determined using a TUNEL Apoptosis kit (cat. no. BA27A; Nanjing Biobox Biotech Co., Ltd.), according to the manufacturer's instructions. After deparaffinization, sections of kidney tissue were decomposed with protease K and successively incubated in terminal transferase buffer with streptavidin-FITC-conjugated antibodies and a POD-conjugated anti-FITC solution, followed by staining with a diaminobenzidine solution (all in the TUNEL kit). Apoptotic cells were observed under a light microscope in five fields of view (magnification, ×400; DM2500; Leica Microsystems GmbH).
4
Cell Cycle and Apoptosis Analysis
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Transfected A375 and A2508 cells were inoculated into 6-well plates and cultured for 48 hours. For cell cycle analysis, cells were collected, washed with PBS, fixed in 75% ethanol, incubated with 50 µg/mL propidium iodide containing 40 µg/mL RNase for 30 minutes, followed by a FACSCalibur flow cytometer (BD Biosciences, San Jose, CA, USA) to measure the distribution of cells in G0/G1, S, and G2/M phases. For apoptosis detection, cells were harvested, washed with PBS, stained with a fluorescein isothiocyanate Annexin V Apoptosis Detection Kit (BD Biosciences), and then subjected to a flow cytometer (BD Biosciences) for the examination of apoptotic rate.
5
Annexin V Apoptosis Assay in BC Cells
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The BC cells were treated with cisplatin and hsa-miR-30a-3p mimic for 24 h, and the apoptotic cells were detected using the fluorescein isothiocyanate Annexin V apoptosis detection kit (BD Biosciences, NJ, USA), according to the manufacturer’s instructions. The populations with Annexin V−/PI−, Annexin V+/PI−, Annexin V+/PI+, and Annexin V−/PI+ cells showed live, early apoptotic, late apoptotic, and necrotic cells that were estimated using the Accuri C5 flow cytometer (BD Biosciences, NJ, USA), and the resulting data were analyzed using CellQuest Pro software (BD Biosciences, NJ, USA).
6
Coculture of MC38 Cells and MPE Macrophages
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MC38 cells were cocultured with MPE macrophages using a cell culture insert with 0.4 μm pores (Corning, NY, USA). MPE macrophages isolated from WT and IL-10−/− mice were seeded within the upper chamber, while MC38 cells (2 × 105 cells/mL) were plated on the lower chamber. Subsequently, the upper chamber was placed onto the plates. After coculture for 24 hr, MC38 cells were collected, and the apoptotic cells were analyzed using a fluorescein isothiocyanate Annexin V Apoptosis Detection Kit (BD Biosciences).
7
Annexin V Apoptosis Assay for Huh7 and HepG2 Cells
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The differences in apoptosis were examined using an Annexin V apoptosis assay. Briefly, Huh7 and HepG2 cells were seeded in 6-well plates at 2 × 105 per well for 48 h under normoxic or hypoxic conditions prior to treatment. After sorafenib treatment/transfection, cells were harvested via trypsinization, washed twice with Phosphate buffered saline (PBS), and stained with a fluorescein isothiocyanate Annexin V Apoptosis Detection kit (BD Pharmingen, Franklin Lakes, NJ, USA). Flow cytometry (Thermo Fisher Scientific, Waltham, Massachusetts, USA) was used to determine the cell apoptosis ratio.
8
Assessing Cell Viability and Apoptosis
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Cell viability was measured using the Cell Viability Assay Kit (EZ-Cytox, DOGEN, Daejeon, Korea). Human CRC HCT116, DLD-1, HT29, and SW620 cells, and normal colon FHC and CCD-18Co cells (1 × 104 cells per well) were seeded on 96-well plates and then treated as described in the Results section. The cells were then incubated with the EZ-Cytox reagent and incubated for 2 h at 37 °C in an atmosphere of 5% CO2. Absorbance at 450 nm was determined using a microplate reader (SPECTRA190, Molecular Devices, Sunnydale, CA, USA).
To detect the translocation of phosphatidylserine, a marker of apoptosis, from the inner to the outer leaflet of the plasma membrane, the cells were stained with Annexin V according to the manufacturer’s protocol for the fluorescein isothiocyanate Annexin V Apoptosis Detection Kit (BD Biosciences, San Diego, CA, USA). Flow cytometry was performed using an Accuri C6 flow cytometer (BD Biosciences).
To detect the translocation of phosphatidylserine, a marker of apoptosis, from the inner to the outer leaflet of the plasma membrane, the cells were stained with Annexin V according to the manufacturer’s protocol for the fluorescein isothiocyanate Annexin V Apoptosis Detection Kit (BD Biosciences, San Diego, CA, USA). Flow cytometry was performed using an Accuri C6 flow cytometer (BD Biosciences).
9
Apoptosis Analysis by Flow Cytometry
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To evaluate the sub-G0 DNA content, cells were stained with propidium iodide (PI, 75 μM ) in PBS containing 0.1% Nonidet P-40. DNA content was determined by flow cytometry using a FACScalibur instrument and CellQuest software (BD Biosciences, San Jose, CA, USA). The percentage of apoptotic cells was also determined with a fluorescein isothiocyanate-Annexin V Apoptosis Detection kit (BD Biosciences) according to the manufacturer’s instructions.
10
Apoptosis Assay with Glycerol and Rapamycin
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Equal numbers of cells transfected with lentiviral vector were cultivated in each well of a 6-well plate. All cells were collected using trypsin solution (without EDTA, Nalgene, Rochester, NY, USA) and stained with a fluorescein isothiocyanate Annexin V Apoptosis Detection kit (BD Pharmingen, Franklin Lakes, NJ, USA). A flow cytometer (BD Biosci-ences, San Jose, CA, USA) was employed to determine the apoptosis ratio of the cells. Glycerol was supplemented at the concentrations indicated above. Rapamycin was added at concentration of 1 µmol/L.28
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