Invitrogen™ eBioscience™ Annexin V Apoptosis Detection Kits

Catalog No.	Quantity	Excitation/Emission	Conjugate
50-927-6	50 Tests	405, 546/450, 647	eFluor 450, 7-AAD
50-927-5	200 Tests	488, 535/520, 617	FITC, Propidium Iodide
50-927-7	200 Tests	405, 546/450, 647	eFluor 450, 7-AAD
50-927-9	200 Tests	650, 535/660, 617	APC, Propidium Iodide
50-928-0	50 Tests	-	PCP-eFluor 710
50-928-1	200 Test	-	PCP-eFluor 710
50-929-7	100 Tests	488, 535/520, 617	FITC, Propidium Iodide
50-929-9	20 Tests	488, 535/520, 617	FITC, Propidium Iodide
50-930-1	300 Tests	488, 535/520, 617	FITC, Propidium Iodide
50-246-343	50 Tests	-	PE, 7-AAD
50-246-344	200 Tests	-	PE, 7-AAD
50-246-345	50 Tests	-	PE/Cy7
50-246-346	200 Tests	-	PE/Cy7
50-929-4	100 Tests	NA, 535/NA, 617	Biotin, Propidium Iodide
50-929-5	20 Tests	NA, 535/NA, 617	Biotin, Propidium Iodide
50-929-6	300 Tests	NA, 535/NA, 617	Biotin, Propidium Iodide
50-112-8958	50 Tests	488, 535/520, 617	FITC, Propidium Iodide
50-112-9048	50 Tests	650, 535/660, 617	APC, Propidium Iodide

Catalog No. 50-927-6 Supplier Invitrogen™ Supplier No. 88800672

Description

Includes

• Cat. No. 88-8006-72:
• 10X Binding Buffer
• Annexin V eFluor™ 450 : 5μL/test, 50 tests Protect from light.
• 7-AAD Viability Staining Solution : 5μL/test, 100 tests Protect from light.
• Cat. No. 88-8006-74:
• 10X Binding Buffer
• Annexin V eFluor™ 450 : 5μL/test, 200 tests Protect from light.
• 7-AAD Viability Staining Solution : 5μL/test, 2x100 tests Protect from light.

Kits utilize viability dye-conjugated annexin V to label phosphatidylserine (PS), an early marker of apoptosis, on the extracellular membrane.

Easily distinguish cells undergoing early and late apoptosis with eBioscience and Annexin V Apoptosis Detection kits for flow cytometry analysis. These kits utilize viability dye-conjugated annexin V to label phosphatidylserine (PS), an early marker of apoptosis, on the extracellular membrane. In late-stage apoptosis, as cell integrity is lost, both annexin V and the viability dye traverse and bind PS on the interior of the plasma membrane.

Annexins are a family of calcium-dependent phospholipid-binding proteins that preferentially bind phosphatidylserine (PS). Under normal physiologic conditions, PS is predominantly located in the inner leaflet of the plasma membrane. Upon initiation of apoptosis, PS loses its asymmetric distribution across the phospholipid bilayer and is translocated to the extracellular membrane leaflet marking cells as targets of phagocytosis. Once on the outer surface of the membrane, PS can be detected by fluorescently labeled annexin V in a calcium-dependent manner.

In early-stage apoptosis, the plasma membrane excludes viability dyes such as propidium iodide (PI), 7-AAD, or fixable viability dyes such as eFluor 450. These cells will stain with annexin V but not a viability dye, thus distinguishing cells in early apoptosis. However, in late-stage apoptosis, the cell membrane loses integrity, thereby allowing annexin V to also access PS in the interior of the cell. A viability dye can be used to resolve these late-stage apoptotic and necrotic cells (annexin V, viability dye-positive) from the early-stage apoptotic cells (annexin V positive, viability dye-negative).

Current viability stains available with the eBioscience Annexin V Apoptosis Detection kits include FITC, eFluor 450, allophycocyanin (APC), phycoerythrin (PE), PE-cyanine7, and biotin.

Specifications

• Description: Annexin V Apoptosis Detection Kit eFluor 450
• Quantity: 50 Tests
• Format: Kit
• Product Type: Apoptosis Detection Kit
• Content And Storage: • Annexin V Binding Buffer (10X), 1 bottle (30 mL)
  • Annexin V eFluor 450, 1 vial (0.25 mL)
  • 7-AAD Viability Staining Solution, 1 vial (0.5 mL)
  
  Note: Fixable Viability Dye eFluor 450 is not recommended for use with Annexin V Apoptosis Detection Kits. 2°C to 8°C. Avoid temperature fluctuations.
• Excitation/Emission: 405, 546/450, 647
• Shipping Condition: Wet Ice
• Product Line: eBioscience
• Conjugate: eFluor 450, 7-AAD
• For Use With (Equipment): Fluorescence Microscope, Flow Cytometer

Frequently Asked Questions (FAQs)

Can I use eFluor 450-conjugated antibodies with eBioscience Annexin V Apoptosis Detection kits?

Yes, eFluor 450-conjugated antibodies are compatible with eBioscience Annexin V Apoptosis Detection kits.

I trypsinized my adherent cells and labeled with annexin V, and now my flow data is showing a high percentage of apoptotic cells even for control, untreated cells. What is the problem?

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).

Can I detect annexin V staining in an imaging assay?

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.

When should I stain adherent cells with annexin V for flow cytometric analysis? Before or after I trypsinize them?

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).

Can I fix my cells after annexin V staining?

Yes, this is possible. We have established protocols for annexin V staining combined with intracellular staining of lymphocytes that can be found here. The most important step is to leave some binding buffer in the suspension when fixation is started. Compared to staining of live cells, the intensity of the annexin V signal may be somewhat reduced.

Safety and Handling

WARNING: Cancer - www.P65Warnings.ca.gov