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Molecular Probes™ Dead Cell Apoptosis Kits with Annexin V for Flow Cytometry
Description
Easily differentiate live, dead, and apoptotic cells during flow cytometry with our Dead Cell Apoptosis kits with Annexin V conjugates including Alexa Fluor 488, FITC, propidium iodide, PE, APC, and SYTOX Green. These conjugates distinguish live, dead, or apoptotic cells via different dyes, which is vital for confirming cell viability and apoptosis using multi-parametric studies.
Superior brightness
Unlike other Annexin V kits that have lower protein concentrations or purification levels, the Annexin V Alexa Fluor 488 conjugate is optimized for flow cytometry to provide the largest separation between apoptotic and live cells. The Alexa Fluor 488 dye is a superior green-fluorescent dye with a spectrum similar to fluorescein (FITC).
High binding efficiency
Annexin V conjugates are made from a highly purified cys-annexin, resulting in higher binding efficiency and extremely accurate characterization of the apoptotic process.
Multi-parametric
Many publications require a minimum of two different ways to identify apoptotic cells. This multi-parametric kit detects phosphatidyl serine (PS) on the cytoplasmic surface of the cell membrane, while membrane integrity is determined via propidium iodide.
Dead Cell Apoptosis Kit with Annexin V Alexa Fluor 488 & PI
The Dead Cell Apoptosis Kit with Annexin V Alexa Fluor 488 & Propidium Iodide (PI) (V13241, V13245) is used in flow cytometry to measure early apoptosis by detecting phosphatidyl serine (PS) expression and membrane permeability. When cells are stained with Annexin V and propidium iodide, apoptotic cells expressing PS show green fluorescence, which can be detected in the FITC channel, and low red fluorescence. Dead or necrotic cells show bright red fluorescence and no green fluorescence, while live cells show no green or red fluorescence.
Dead Cell Apoptosis Kit with Annexin V PE and SYTOX Green
The Dead Cell Apoptosis Kit with Annexin V PE and SYTOX Green detects PS externalization in apoptotic cells during flow cytometry using recombinant annexin V conjugated to the orange fluorescent phycobiliprotein R-PE, while dead cells are detected with SYTOX Green nucleic acid stain. After treatment with both probes, apoptotic cells show orange fluorescence, dead cells show green fluorescence, and live cells show little or no fluorescence. These populations can easily be distinguished in the 530/30 nm and 585/42 nm bandpass filters with a 488 nm laser flow cytometer.
Dead Cell Apoptosis Kit with Annexin V APC and SYTOX Green
The Dead Cell Apoptosis Kit with Annexin V APC and SYTOX Green detects PS externalization in apoptotic cells during flow cytometry using recombinant annexin V conjugated to red laser-excited allophycocyanin, and dead cells using SYTOX Green nucleic acid stain. Apoptotic cells are detected by annexin V binding to externalized PS. Live cells show little or no fluorescence, apoptotic cells show red fluorescence and very little green fluorescence, and late apoptotic cells show a higher level of red and orange fluorescence. These populations can easily be distinguished using a flow cytometer that has both 488 nm and 633 nm excitation sources (an argon-ion laser and a HeNe laser).
The Dead Cell Apoptosis Kit with Annexin V FITC and PI
The Dead Cell Apoptosis Kit with Annexin V FITC and PI detects PS externalization in apoptotic cells using recombinant annexin V conjugated to green-fluorescent FITC dye and dead cells using PI. Necrotic cells that stain with PI show red fluorescence. After treatment with both probes, apoptotic cells show green fluorescence, dead cells show red and green fluorescence, and live cells show little or no fluorescence.
Specifications
Specifications
| Description | Dead Cell Apoptosis Kit with Annexin V APC and SYTOX Green, for flow cytometry |
| Quantity | 1 Kit |
| Kit Contents | Contains 1 vial of annexin V, APC conjugate (250 μL), 1 vial of SYTOX green stain (100 μL),and 1 bottle of annexin binding buffer (5X solution, 15 mL) . |
| Product Type | Apoptosis Detection Kit |
| Content And Storage | Store in refrigerator (2°C to 8°C) and protect from light. |
| Flow Cytometer Laser Lines | 633/635, 488 |
| Excitation/Emission | 503, 650/524, 660 |
| No. of Reactions | 50 |
| Shipping Condition | Wet Ice |
| For Use With (Equipment) | Fluorescence Microscope, Flow Cytometer |
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Frequently Asked Questions (FAQs)
The binding mode of SYTO nucleic acid stains is unknown. However, the behavior of these and related nucleic acid dyes suggests the following binding properties:
1.They appear to contact the solvent (suggested by sensitivity to salt, divalent cations, and in particular, SDS) and thus are likely to have contacts in the grooves.
2.All SYTO dyes appear to show some base selectivity and are thus likely to have minor groove contacts.
3.They can be removed from nucleic acid via ethanol precipitation; this characteristic is not shared by ethidium bromide and other intercalators. Likewise, the dyes are not removed from nucleic acid via butanol or chloroform extraction. These extraction methods do remove ethidium bromide from nucleic acid.
4. SYTO binding is not affected by nonionic detergents.
5. SYTO dyes are not quenched by BrdU, so they do not bind nucleic acids in precisely the same way as Hoechst 33342 and DAPI ((4′,6-diamidino-2-phenylindole).
SYBR Green I has shown little mutagenicity on frameshift indicator strains, indicating that it isn't likely to strongly intercalate.
These products are cells that died without undergoing apoptosis. A PI+ only population could be either a result of experimental condition or sample preparation.
Trypsinization or mechanical scraping of cells temporarily disrupts the plasma membrane, allowing annexin V to bind phosphatidylserine on the cytoplasmic surface of the cell membrane and thus leading to false positive staining. Allow the cells to recover for about 30 minutes in optimal cell culture conditions and medium after trypsinizing/scraping so that they can recover their membrane integrity before staining. For lightly adherent cell lines, such as HeLa and NIH 3T3, another option is to use non-enzyme treatments like Gibco Cell Dissociation Buffer (Cat. No. 13151014).
Annexin V staining is not typically used in imaging experiments; it is a better reagent for flow cytometry analysis. All cells will stain to some extent, so it can be difficult to distinguish a relatively bright annexin V-stained cell from a dimmer non-apoptotic cell. Caspase activation, detected using our CellEvent Caspase 3/7 or Image-iT LIVE Caspase detection kits, is a better method for detecting apoptosis in an imaging assay.
Trypsinize first and then allow the cells to recover about 30 minutes in optimal cell culture conditions and medium before staining with annexin V conjugates. Trypsinization or mechanical scraping of cells temporarily disrupts the plasma membrane, allowing for annexin V to bind phosphatidylserine on the cytoplasmic surface of the cell membrane and thus leading to false positive staining. For lightly adherent cell lines such as HeLa and NIH 3T3, you could use a less harsh (non-enzymatic) dissociation product like Gibco Cell Dissociation Buffer (Cat. No. 13151014).
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