After the indicated treatment with palmitate and Ang (1–7), cells were harvested and the rate of apoptosis was determined by flow cytometry with an Annexin V-FITC/PI assay (BD Pharmingen, Franklin Lakes, NJ) according to the manufacturer's instructions.
Annexin 5 fitc pi assay
Manufactured by BD
Sourced in United States
The Annexin V-FITC/PI assay is a laboratory analysis tool used to detect and quantify apoptosis, or programmed cell death, in cell samples. The assay utilizes Annexin V, a protein that binds to phosphatidylserine, and propidium iodide (PI), a fluorescent dye that intercalates with DNA, to distinguish between viable, apoptotic, and necrotic cells.
Automatically generated - may contain errors
6 protocols using annexin 5 fitc pi assay
1
Apoptosis Measurement by Flow Cytometry
2
Apoptosis Regulation in LPS-Stimulated HUVECs
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After incubating HUVECs with 100 ng/mL LPS for 24 h, we further incubated the cells with GP + EM and TK-MLP@(GP + EM) NPs for an additional 24 h. The apoptosis of HUVECs was assessed using flow cytometry with an Annexin V-FITC/PI assay, following the manufacturer’s instructions (BD, CA, USA).
Subsequently, the cells were lysed, and the protein samples were subjected to SDS-PAGE gel separation. The separated proteins were then transferred onto a polyvinylidene fluoride (PVDF) membrane. The membranes were blocked with TBST containing 5% skim milk and incubated overnight at 4℃ with primary antibodies against Cleavedcaspase-3, Bcl-2, BAX, and β-actin. Following incubation, the membranes were washed and incubated with a secondary antibody at room temperature for 2 h. Finally, the membranes were washed and visualized using the BIO-RAD ChemiDoc XRS chemiluminescence system.
Subsequently, the cells were lysed, and the protein samples were subjected to SDS-PAGE gel separation. The separated proteins were then transferred onto a polyvinylidene fluoride (PVDF) membrane. The membranes were blocked with TBST containing 5% skim milk and incubated overnight at 4℃ with primary antibodies against Cleavedcaspase-3, Bcl-2, BAX, and β-actin. Following incubation, the membranes were washed and incubated with a secondary antibody at room temperature for 2 h. Finally, the membranes were washed and visualized using the BIO-RAD ChemiDoc XRS chemiluminescence system.
3
Apoptosis Quantification in MSB-1 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Apoptotic morphology: for morphological examination, distinct treated MSB-1 cells were stained with 4′,6′-diamidino-2-phenylindole (DAPI; Sigma-Aldrich), and those with fragmented or condensed nuclei in deep staining were counted as apoptotic cells.
Annexin V-fluorescein isothiocyanate/propidium iodide (FITC/PI) assay: for the treated cells, an annexin V-FITC/PI assay (BD Biosciences Pharmingen) was performed according to the manufacturer’s protocol; after staining, cells were analyzed by a FACSCalibur instrument (Becton Dickinson, San Jose, CA). Experiments were repeated at least three times in duplicates.
Annexin V-fluorescein isothiocyanate/propidium iodide (FITC/PI) assay: for the treated cells, an annexin V-FITC/PI assay (BD Biosciences Pharmingen) was performed according to the manufacturer’s protocol; after staining, cells were analyzed by a FACSCalibur instrument (Becton Dickinson, San Jose, CA). Experiments were repeated at least three times in duplicates.
4
Annexin V-FITC/PI Apoptosis Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Apoptosis was evaluated with an annexin V-FITC/PI assay (BD Biosciences Pharmingen). Marc-145 cells were challenged with CH-1a (MOI =0.01) in the presence of CP1 (480 μg/ml) or YPD (control) for 24, 48, or 72 h. Apoptosis was then measured with annexin V-FITC/PI staining, according to the manufacturer’s protocol. The cells were then analyzed with fluorescence microscopy (Carl Zeiss).
5
Endothelial Cell Apoptosis and Permeability
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
MAEC were pretreated with or without rMYDGF (50 mg/ml) for 48 hours, and an additional treatment with or without PA (0.4 mM for 16 hours; Sigma-Aldrich, USA) was done when needed. Cell apoptosis was determined by flow cytometry with an annexin V–FITC/PI assay (BD, CA, USA) according to the manufacturer’s instructions (11 (link)). The endothelial permeability was measured by the In Vitro Vascular Permeability Assay Kit (Millipore) according to the manufacturer’s instructions.
6
Evaluating Apoptosis in DF-1 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Apoptosis was evaluated by apoptotic morphology and Annexin V-fluorescein isothiocyanate/propidium iodide (FITC/PI) assay for which cells were treated in similar ways as for the cell proliferation assay. About 24 hours after transfection with gga-miR-375 or miR-NC, DF-1 cells (1.0 × 105 per millilitre) were seeded respectively into a 6-well plate and incubated for another 24, 48, or 72 hours under serum starvation; a blank control was also used. Then, Annexin V-FITC/PI assay (BD Biosciences Pharmingen, Franklin Lakes, NJ, USA) was performed according to the manufacturer's protocol. After staining, cells were analysed by FACS Calibur (Becton Dickinson, San Jose, CA, USA). For morphologic examination, after 48 hour serum starvation treatment, cells were stained with 4′-6′-diamidino-2-phenylindole (DAPI; Sigma-Aldrich Co, St Louis, MO, USA) and those with fragmented or condensed nuclei in deep staining were counted as apoptotic cells. At least 500 cells were counted for each plate. The background luminescence associated with cell culture and assay reagent (blank reaction) was subtracted from the experimental value.
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!