Apoptosis induced by SIE was measured by Annexin-V-FITC-PI staining using flow cytometer. MCF-7 cells (1 × 106/well) were seeded in 6-well plate and allowed to grow for 24 h. Cells were treated with different concentration of SIE for 72 h. At the end of the treatment, all cells including floating cells were harvested, washed with PBS, and stained with Annexin-V-FITC and propidium iodide (Sigma-Aldrich) for 10 min at RT [68 (link), 69 (link)]. Samples were acquired by flow cytometer FACS caliber (BD biosciences).
Annexin 5 fitc and propidium iodide
Manufactured by Merck Group
Sourced in Germany
Annexin V-FITC and propidium iodide (PI) are fluorescent labeling reagents commonly used in flow cytometry and microscopy applications. Annexin V binds to phosphatidylserine, a phospholipid that is externalized during apoptosis, while PI stains DNA in cells with compromised cell membranes. These reagents can be used to detect and quantify apoptotic and necrotic cells.
Automatically generated - may contain errors
Lab products found in correlation
10 protocols using annexin 5 fitc and propidium iodide
1
Apoptosis Measurement by Annexin-V-FITC-PI Staining
2
Quantifying Apoptosis and Necrosis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were detached from the petri dishes by trypsinization in order to minimize cell re-aggregation and permit further cytometric studies, as we previously described for primary cultured hepatocytes [7 (link)], with slight modifications. After gently homogenization in the culture medium/PBS and harvest (5 min, 400 g), 100,000 cells were carefully re-suspended in the appropriate buffer. Apoptotic externalization of phosphatidylserine and cell death was assessed by staining with Annexin V-FITC and propidium iodide (Sigma Chemical Co.) coupled to flow cytometric analysis (Cell Sorter BD FACSAria II, BD Biosciences), following the manufacturers' instructions. Detection of green and red fluorescence was performed, and the proportion of Annexin V (apoptotic) and propidium iodide (necrotic –by the treatment and/or manipulation–) positive cells were determined in the indicated experimental groups. Green and red fluorescence intensities detected in non stained cells were used to set the thresholds for each channel.
3
Apoptosis Analysis of T47D and MB231 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After transfection, T47D and MB231 cells resuspended in binding buffer were doubled-stained with 10 μL of Annexin V-FITC and propidium iodide (Sigma-Aldrich). The apoptotic cells were determined by a FACScan flow cytometer.
4
Apoptosis Analysis of SH-SY5H Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After transfection with shRNA for 24 h, SH-SY5H cells were collected and resuspended at a concentration of 1 × 10 7 cells/ml. The cell suspension was stained with Annexin-V-FITC and propidium iodide (Sigma-Aldrich) for 15 minutes. Apoptosis was detected using a FACS Calibur flow cytometer (BD Biosciences, San Jose, CA, USA). Cells (2 ×10 4 ) were collected, recorded, and the data were presented as twoparameter dot-plots.
5
Annexin V-FITC/PI Cell Apoptosis Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After treated with L‐carnitine, cells were washed twice with PBS and then stained according to Annexin V‐FITC/PI kit protocol (BD PharMingen). Briefly, cells were incubated with Annexin V‐FITC and propidium iodide (PI; Sigma‐Aldrich) and incubated for 15 min in the dark. Samples were read on a FACScan flow cytometer (Becton Dickinson).
After treated with L‐carnitine, cells were treated with 0.25% trypsin (Sigma‐Aldrich) and detached from the plate. Cells were stained with propidium oxide (BD Biosciences). The distribution of the cells in G0/G1, S, and G2/M phases were determined with FACScan cytometry (Becton Dickinson).
After treated with L‐carnitine, cells were treated with 0.25% trypsin (Sigma‐Aldrich) and detached from the plate. Cells were stained with propidium oxide (BD Biosciences). The distribution of the cells in G0/G1, S, and G2/M phases were determined with FACScan cytometry (Becton Dickinson).
6
Apoptosis and Cell Cycle Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were washed using PBS. And the cell pellets were resuspended and stained with annexin V-FITC and propidium iodide (PI) (Sigma‑Aldrich). The rate of cell apoptosis was analyzed using a FACsorter (BD Biosciences, San Jose, CA, USA) after incubating 15 min at room temperature. For cell cycle detection, cells were fixed with precooled 70% ethanol overnight at -20°C. After centrifugation, the cells were resuspended with RNase solution in a 37°C water bath for 30 min. Then propidium iodide staining solution was added and incubated for 30 min in the dark at room temperature. The cell cycle distribution was determined using a FACsorter.
7
Apoptosis and Cell Cycle Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells with designated treatments were harvested, washed, and stained with Annexin V-FITC and propidium iodide (PI) (Sigma-Aldrich), according to the manufacturer’s protocols. Cells were then analyzed by the BD FACSCalibur flow cytometer (BD Biosciences) using a Cell Quest software (version 5.0; BD Biosciences). For analyzing cell cycle distribution, cells were only stained with PI.
8
Apoptosis and Cell Cycle Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were washed using PBS. And the cell pellets were resuspended and stained with annexin V-FITC and propidium iodide (PI) (Sigma-Aldrich). The rate of cell apoptosis was analyzed using a FACsorter (BD Biosciences, San Jose, CA, USA) after incubating 15 min at room temperature. For cell cycle detection, cells were fixed with precooled 70% ethanol overnight at -20°C. After centrifugation, the cells were resuspended with RNase solution in a 37°C water bath for 30 min. Then propidium iodide staining solution was added and incubated for 30 min in the dark at room temperature. The cell cycle distribution was determined using a FACsorter.
9
Cytotoxicity Assessment via Flow Cytometry and Fluorescence Microscopy
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cell death was assessed by staining the cells with AnnexinV-FITC and propidium iodide (PI) (Sigma-Aldrich, Steinheim, Germany) according to the manufacturer’s instructions (eBioscience, San Diego, CA, USA) and subsequent analysis by flow cytometry. MCF-7 cells (5.0 × 104 cells/cm2) were seeded in triplicate in 24-wells culture and after an overnight incubation, washed with fresh medium and incubated for 24 h with the compounds or vehicle alone (control cells) in DMEM. After trypsinization, 1.0 × 106 cells/mL in combining buffer were incubated with Annexin V-FITC and PI solution at room temperature in the dark for 15 min. Then samples of at least 1.0 × 104 cells were subjected to FACS analysis using appropriate 2-bidimensional gating method.
Acridine orange/ethidium bromide (AO/EB) fluorescence staining was also used to identify cell apoptosis. After 24 h treatment, the cells were washed with PBS and stained with AO/EB solution (100 μg/mL, 1:1). After 20 s the AO/EB solution was discarded and cells were immediately visualized by fluorescence microscopy (Leica; Wetzlar, Germany). Multiple photos were taken at randomly-selected areas of the well to obtain representative data.
Acridine orange/ethidium bromide (AO/EB) fluorescence staining was also used to identify cell apoptosis. After 24 h treatment, the cells were washed with PBS and stained with AO/EB solution (100 μg/mL, 1:1). After 20 s the AO/EB solution was discarded and cells were immediately visualized by fluorescence microscopy (Leica; Wetzlar, Germany). Multiple photos were taken at randomly-selected areas of the well to obtain representative data.
10
Apoptosis and Cell Cycle Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were washed using PBS. And the cell pellets were resuspended and stained with annexin V-FITC and propidium iodide (PI) (Sigma-Aldrich). The rate of cell apoptosis was analyzed using a FACsorter (BD Biosciences, San Jose, CA, USA) after incubation for 15 min at room temperature. For cell cycle detection, cells were fixed with pre-cooled 70% ethanol overnight at -20 °C. After centrifugation, the cells were resuspended with RNase solution in a 37 °C water bath for 30 min. Then propidium iodide staining solution was added and incubated for 30 min in the dark at room temperature. The cell cycle distribution was determined using a FACsorter. And for the transfection efficiency also was tested by flow cytometric. Each experiment was repeated at least three times.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!