In normal cells, phosphatidylserines (PS, membrane phospholipids) are held on the inner layer of the cell membrane, so Annexin V does not attach to the cells. During early apoptosis, the PS are exposed on the outer layer, where they attach to the FITC-labeled Annexin V and stain the cell surface green. During late apoptosis, propidium iodide (PI) enters the cell and stains the contents red.
Jurkat cells were apoptosis induced with staurosporine with its concentration of 1 µg/ml at 37 °C for 3.5 hours and then observed under a fluorescent microscope.
FITC-labeled Annexin V (Green) / PI (Red)
Jurkat cells were apoptosis induced with staurosporine (1 µg/ml) at 37 °C for 3.5 hours and then analyzed with a flow cytometer.
|Reagents||for 100 assays|
|Annexin V-FITC Conjugate||250 µl||2|
|PI Solution||250 µl||2|
|Annexin V Binding Buffer (10x)||10 ml||2|
Cell type: Mouse B16 melanoma cells
Number of cells: 10,000
Instrument: Gallios (Beckman Coulter Inc.)
Apoptosis was induced by cycloheximide (1 µg/ml) or UV irradiation at 37°C for 3.5 hours. Cells were analyzed by flow cytometry. Our product showed about the same performance as the product from company A. The control cells did not display any signs of apoptosis. Treatment with cycloheximide resulted in many cells in early apoptosis. UV irradiation also induced apoptosis in the tested cells.
--: FITC-labeled Annexin V and PI both had low fluorescence values. Live cells.
+-: FITC-labeled Annexin V had high fluorescence value, PI was low. Early apoptosis.
++: FITC-labeled Annexin V and PI both had high fluorescence values. Late apoptosis.
Data Courtesy of Assistant professor Yuuki Takahashi , Graduate School and Faculty of Pharmaceutical Sciences, Kyoto University
|Control (untreated)||Starvation (48 hours)|
Apoptotsis was induced by removing glucose and starving the cells for 48 hours. Upon analysis using an imaging cytometer, some cells were observed in early apoptosis.
Data Courtesy of Assistant Professor Hitoshi Gotoh Department of Biology / Developmental Neurobiology, Liberal Arts and Sciences, Department of Biology, Kyoto Prefectural University of Medicine
Dilute Annexin V Binding Buffer (10x) 10-fold with distilled water.
*Although adherent cells are not frequently used for Annexin V, FITC flow cytometric analyses to avoid cell membrane damage from the cell detachment process, Casiola-Rosen et al. and van Engelend et al. have reported methods on utilizing Annexin V for flow cytometry with adherent cell types.
|excitation / emission|
|Annexin V-FITC||494 nm / 518 nm|
|PI||535 nm / 617 nm|
|Annexin V-FITC Apoptosis Detection Kit||15342-54||4℃||100 tests||473.00||Buy|