Cell Meter™ Annexin V Binding Apoptosis Assay Kit *Green Fluorescence Optimized for Flow Cytometry*
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Telephone | 1-800-990-8053 |
Fax | 1-800-609-2943 |
sales@aatbio.com | |
International | See distributors |
Bulk request | Inquire |
Custom size | Inquire |
Shipping | Standard overnight for United States, inquire for international |
Correction Factor (260 nm) | 0.21 |
Correction Factor (280 nm) | 0.11 |
Extinction coefficient (cm -1 M -1) | 750001 |
Excitation (nm) | 491 |
Emission (nm) | 516 |
Quantum yield | 0.91 |
H-phrase | H303, H313, H333 |
Hazard symbol | XN |
Intended use | Research Use Only (RUO) |
R-phrase | R20, R21, R22 |
UNSPSC | 12352200 |
Overview | ![]() ![]() |
Correction Factor (260 nm) 0.21 | Correction Factor (280 nm) 0.11 | Extinction coefficient (cm -1 M -1) 750001 | Excitation (nm) 491 | Emission (nm) 516 | Quantum yield 0.91 |
Platform
Flow cytometer
Excitation | 488 nm laser |
Emission | 530/30 nm filter |
Instrument specification(s) | FITC channel |
Components
Example protocol
AT A GLANCE
Prepare cells with test compounds (200 µL/sample).
Add Annexin V-iFluor® 488 stock solution.
Incubate at room temperature for 30 - 60 minutes.
Analyze cells using a flow cytometer with a 530/30 nm filter (FITC channel).
Before starting the experiment, thaw the vial of 100X Propidium Iodide (Component C) at room temperature.
CELL PREPARATION
For guidelines on cell sample preparation, please visit:
https://www.aatbio.com/resources/guides/cell-sample-preparation.html
SAMPLE EXPERIMENTAL PROTOCOL
Treat cells with test compounds for a desired period of time (4 - 6 hours for Jurkat cells treated with camptothecin) to induce apoptosis.
Note: Annexin V flow cytometric analysis on adherent cells is not routinely tested since specific membrane damage may occur during cell detachment or harvesting. However, methods for utilizing Annexin V for flow cytometry on adherent cell types have been previously reported by Casiola-Rosen et al. and van Engelend et al.
Centrifuge the cells to get 1 - 5 × 105 cells/tube.
Resuspend cells in 200 µL of Assay Buffer (Component B).
Add 2 µL of Annexin V-iFluor® 488 (Component A) into the cells.
Optional: Add 2 µL of 100X Propidium Iodide (Component C) for necrosis cells.
Incubate at room temperature for 30 to 60 minutes, protected from light.
Optional: Add 200 to 300 µL of Assay Buffer (Component B) to increase volume before analyzing the cells with a flow cytometer.
Monitor the fluorescence intensity of Annexin V-iFluor® 488 using a flow cytometer with a 530/30 nm filter (FITC channel). Measure the cell viability using the 610/20 nm filter (PE-Texas Red channel) after adding propidium iodide to the cells.
Spectrum
![spectrum](/_next/image?url=https%3A%2F%2Fimages.aatbio.com%2Fspectra%2Fifluor_488.png&w=2048&q=50)
Spectral properties
Correction Factor (260 nm) | 0.21 |
Correction Factor (280 nm) | 0.11 |
Extinction coefficient (cm -1 M -1) | 750001 |
Excitation (nm) | 491 |
Emission (nm) | 516 |
Quantum yield | 0.91 |
Product Family
Name | Excitation (nm) | Emission (nm) | Extinction coefficient (cm -1 M -1) | Quantum yield | Correction Factor (260 nm) | Correction Factor (280 nm) |
Cell Meter™ Annexin V Binding Apoptosis Assay Kit *Orange Fluorescence Optimized for Flow Cytometry* | 557 | 570 | 1000001 | 0.641 | 0.23 | 0.14 |
Cell Meter™ Annexin V Binding Apoptosis Assay Kit *Red Fluorescence Optimized for Flow Cytometry* | 587 | 603 | 2000001 | 0.531 | 0.05 | 0.04 |
Cell Meter™ Annexin V Binding Apoptosis Assay Kit *Deep Red Fluorescence Optimized for Flow Cytometry* | 656 | 670 | 2500001 | 0.251 | 0.03 | 0.03 |
Images
![The detection of binding activity of Annexin V-iFluor® 488 and phosphatidylserine in Jurkat cells. Jurkat cells were treated without (Blue) or with 20 µM camptothecin (Red) in a 37 °C, 5% CO2 incubator for 4-5 hours, and then dye loaded with Annexin V-iFluor® 488 for 30 minutes. The fluorescence intensity of Annexin V-iFluor® 488 was measured with a FACSCalibur (Becton Dickinson) flow cytometer using the FL1 channel.](/_next/image?url=https%3A%2F%2Fimages.aatbio.com%2Fproducts%2Ffigures-and-data%2Fcell-meter-annexin-v-binding-apoptosis-assay-kit-green-fluorescence-optimized-for-flow-cytometry%2Ffigure-for-cell-meter-annexin-v-binding-apoptosis-assay-kit-green-fluorescence-optimized-for-flow-cytometry_INjw1.jpg&w=3840&q=75)
![Effect of FSS on apoptosis and necrosis in tubular cells. Confluent monolayers of HK-2 cells were submitted to FSS 0 (static) or FSS 0.5 Pa (FSS 0.5) for 48h. A/ Cells were stained with Annexin-V and then immediately subjected to analysis of phosphatidylserine externalization (Annexin-V fluorescence, X-axis) and Propidium Iodure (PI) uptake (PI fluorescence, Y-axis) using flow cytometry. Living, early apoptotic or necrotic (primary or secondary) cells were distinguished by the criteria of Annexin-V−/PI−(bottom left quadrant), Annexin-V+/PI− (bottom right quadrant) and Annexin-V+/PI+ (upper right quadrant), respectively. B/ Proportions of early apoptosis and necrosis cells were quantified and results are expressed as a percentage of the total population of cells. Data represent mean ± SEM of 7 experiments. *HK-2 cells were assessed for apoptosis and necrosis using Cell Meter Annexin V Binding Apoptosis Assay Kit (AAT Bioquest), according to the manufacturer's instructions. Source: Graph from <strong>Shear Stress-Induced Alteration of Epithelial Organization in Human Renal Tubular Cells</strong> by Damien Maggiorani, et al., <em>PLoS ONE</em>, July 2015.](/_next/image?url=https%3A%2F%2Fimages.aatbio.com%2Fproducts%2Ffigures-and-data%2Fcell-meter-annexin-v-binding-apoptosis-assay-kit-green-fluorescence-optimized-for-flow-cytometry%2Ffigure-for-cell-meter-annexin-v-binding-apoptosis-assay-kit-green-fluorescence-optimized-for-flow-cytometry_yplww.png&w=3840&q=75)
Citations
Authors: Zeng, Weiquan and Wu, Meizhu and Cheng, Ying and Liu, Liya and Han, Yuying and Xie, Qiurong and Li, Jiapeng and Wei, Lihui and Fang, Yi and Chen, Youqin and others,
Journal: Plos one (2022): e0279851
Authors: Lala, Trisha and Doan, Juleva K and Takatsu, Hiroyuki and Hartzell, H Criss and Shin, Hye-Won and Hall, Randy A
Journal: Journal of Biological Chemistry (2022): 102685
Authors: Yildirim, Merve and Kayalar, Ozgecan and Atahan, Ersan and Oztay, Fusun
Journal: Journal of Biochemical and Molecular Toxicology (2022): e23074
Authors: Shen, Pei-Wen and Chou, Yu-Mei and Li, Chia-Ling and Liao, En-Chih and Huang, Hung-Sen and Yin, Chun-Hao and Chen, Chien-Liang and Yu, Sheng-Jie
Journal: Oncology letters (2021): 1--11
Authors: de Le{\'o}n, Andr{\'e}s and Gibon, Julien and Barker, Philip A
Journal: Eneuro (2020)
Authors: El, D Hajj Diab and Clerc, P and Serhan, N and Fourmy, D and Gigoux, V
Journal: Nanomaterials (Basel, Switzerland) (2018)
Authors: Lin, Yung-Kai and Sharma, Ruchi and Ma, Hsu and Chen, Wen-Shyan and Yao, Chao-Ling
Journal: Journal of the Taiwan Institute of Chemical Engineers (2017)
Authors: Do, Duyen Thi and Phan, Nam Nhut and Wang, Chih-Yang and Sun, Zhengda and Lin, Yen-Chang
Journal: Cytotechnology (2016): 2589--2604
Authors: Maggiorani, Damien and Dissard, Romain and Belloy, Marcy and Saulnier-Blache, Jean-S{\'e}bastien and Casemayou, Audrey and Ducasse, Laure and Gr{\`e}s, Sandra and Belli{\`e}re, Julie and Caubet, C{\'e}cile and Bascands, Jean-Loup and others,
Journal: PLoS One (2015): e0131416
Authors: Maggiorani, Damien and Dissard, Romain and Belloy, Marcy and Saulnier-Blache, Jean-Sébastien and Casemayou, Audrey and Ducasse, Laure and Grès, S and ra , undefined and Bellière, Julie and Caubet, Cécile and Basc, undefined and s, Jean-Loup and others, undefined
Journal: PloS one (2015): e0131416
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