Human keratinocytes (HaCaT cells), one of the human epithelial cells, was used as the representative host cells in the present study (31 (link)). To test the cytotoxicity of aBL to normal human epithelial cells, suspensions of HaCaT cells (106 cells mL−1) were exposed to 405-nm aBL at the irradiance of 108 and 216 J cm−2 in 35-mm petri dishes, respectively. The viability of the cells was determined by flow cytometry after aBL exposure. Annexin V/Propidium Iodide (PI) Apoptosis Kit (Invitrogen; Thermo Fisher) was used to determine if cells were viable (32 ). The confluent cells after the incubation were collected and washed once in cold DPBS, and then resuspended in 1× annexin-binding buffer. Each 100 μL of the cell suspension was incubated with 5 μL of FITC annexin V and 1 μL of 100 μg mL−1 PI working solution at room temperature for 15 min. Immediately after the incubation, 400 μL of 1× annexin-binding buffer was added and the stained cells were analyzed using flow cytometry (Fortessa X-20; BD Biosciences, Franklin Lakes, NJ). The fluorescence emissions at 530 nm and 610 nm were measured.
Annexin 5 propidium iodide apoptosis kit
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Annexin V/Propidium Iodide (PI) Apoptosis Kit is a laboratory reagent used for the detection and quantification of apoptosis in cell samples. It utilizes the binding properties of Annexin V, a calcium-dependent phospholipid-binding protein, and the DNA-binding dye Propidium Iodide to identify cells in various stages of apoptosis.
Automatically generated - may contain errors
Lab products found in correlation
Cell culture supplies, by Thermo Fisher Scientific (2 mentions)
Fortessa x20, by BD (1 mentions)
Osthole, by Merck Group (1 mentions)
Rhodamine 123, by Merck Group (1 mentions)
Cell counting kit 8 cck 8, by Merck Group (1 mentions)
Hoechst 33342, by Merck Group (1 mentions)
Facsdiva software version 6, by BD (1 mentions)
Facscanto 2, by BD (1 mentions)
Cell counting kit 8 cck 8, by Beyotime (1 mentions)
Z vad fmk, by Beyotime (1 mentions)
N acetyl cysteine (nac), by Beyotime (1 mentions)
Ly294002, by Beyotime (1 mentions)
Glutathione (gsh), by Beyotime (1 mentions)
7 protocols using annexin 5 propidium iodide apoptosis kit
1
Evaluating Cytotoxicity of 405-nm aBL on HaCaT Cells
2
Osthole Cytotoxicity and Apoptosis Assay
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Osthole was purchased from Sigma-Aldrich; Merck KGaA (Darmstadt, Germany) (Fig. 1A ), dissolved in dimethyl sulfoxide (DMSO), and stored at −20°C. The final DMSO concentration used was <0.1%. The cell counting kit-8 (CCK-8), Hoechst 33342, and rhodamine 123 were purchased from Sigma-Aldrich; Merck KGaA. The Annexin V/propidium iodide (PI) apoptosis kit was purchased from Invitrogen; Thermo Fisher Scientific, Inc. (Waltham, MA, USA). All antibodies were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). All cell culture supplies were obtained from Gibco; Thermo Fisher Scientific, Inc.
3
Apoptosis Detection by Flow Cytometry
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The cells (1×105/well) were cultured in 6-well plates. An Annexin V/propidium iodide (PI) apoptosis kit (Invitrogen; Thermo Fisher Scientific, Inc.) was used to detect apoptosis. According to the manufacturer's protocol, Annexin-V and PI staining was analyzed using analyzed using a flow cytometer (BD FACSCanto II) with FACSDiva software version 6.1 (both BD Biosciences).
4
Liensinine Cytotoxicity and Apoptosis Evaluation
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Liensinine was purchased from Shanghai Jinsui Bio-Technology Co., Ltd.; the purity was at least 99% as determined by high-performance liquid chromatography. Liensinine was dissolved in dimethyl sulfoxide (DMSO) and stored at -20 ℃. The final liensinine concentration did not exceed 0.1%. Cell Counting Kit-8 (CCK-8) was purchased from Beyotime Institute of Biotechnology Company (Suzhou, Jiangsu, China). Annexin V/Propidium Iodide (PI) Apoptosis Kit was purchased from Invitrogen (Carlsbad, CA, USA). All antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). All cell culture supplies were obtained from Invitrogen (Carlsbad, CA, USA).
5
Quantifying Chondrocyte Apoptosis by Flow Cytometry
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A flow cytometry assay was performed using the annexin V-propidium iodide (PI) apoptosis kit according to the manufacturer’s instructions (Invitrogen, Carlsbad, CA, USA). Chondrocytes were seeded in 6-well plates (1×106 cells/well) and cultured for 48 hours. The cells were then harvested by trypsinization and washed with cold phosphate-buffered saline (PBS), and then resuspended in binding buffer. Annexin V-fluorescein isothiocyanate (FITC) and PI were added to the suspension. The suspension was incubated for 15 min in the dark at room temperature, and then the binding buffer was added. Finally, the samples were analyzed by flow cytometry.
6
Cytotoxicity Evaluation of DSN Compound
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DSN was purchased from Sigma-Aldrich (St. Louis, MO, USA); the purity was at least 98%, as determined by high performance liquid chromatography. DSN was dissolved in dimethyl sulfoxide (DMSO) in a 100 mM stock solution and stored at -20°C. The final DMSO concentration was less than 0.1%. Cell Counting Kit-8 (CCK-8), Hoechst 33342 and Rhodamine 123 kits were purchased from Sigma-Aldrich. Annexin V/Propidium Iodide (PI) Apoptosis Kit was purchased from Invitrogen (Carlsbad, CA, USA). A pan-caspase inhibitor (Z-VAD-FMK) and PI3K inhibitor (LY294002) as well as GSH, NAC and GSH/GSSH and ROS detection kits were purchased from Beyotime Institute of Biotechnology Company (Suzhou, Jiangsu, China). All antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). All cell culture supplies were obtained from Invitrogen (Carlsbad, CA, USA).
7
Apoptosis Quantification in Chondrocytes
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A flow cytometry assay was performed using the annexin V-propidium iodide (PI) apoptosis kit according to the manufacturer's instructions (Invitrogen, Carlsbad, CA, USA). Chondrocytes were seeded in 6-well plates (1 × 106 cells/well) and cultured for 48 h. Then, cells were harvested by trypsinization, washed with cold phosphate-buffered saline (PBS), and resuspended in binding buffer. Annexin V-fluorescein isothiocyanate (FITC) and PI were added to the suspension. The suspension was incubated for 15 min in the dark at room temperature, and then, the binding buffer was added. Finally, the samples were analyzed by flow cytometry.
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