Shop All Apoptosis Kits

Vybrant™ FAM Caspase-3 and -7 Assay Kit, for flow cytometry (Invitrogen™)

The Vybrant™ FAM Caspase-3 and -7 Assay Kit employs a novel approach to detect active caspases that is based on a fluorescent inhibitor of caspases (FLICA™) methodology, essentially an affinity label. The reagent associates a fluoromethyl ketone (FMK) moiety, which can react covalently with a cysteine, with a caspase-specific amine acid sequence. For caspase-3 and -7, this recognition sequence is aspartic acid-glutamic acid-valine-aspartic acid (DEVD). A fluorescein group is attached as a reporter. The FLICA reagent is thought to interact with the enzymatic reactive center of an activated caspase via the recognition sequence, and then to attach covalently through the FMK moiety. The FLICA inhibitor is cell permeant and noncytotoxic. Unbound FLICA molecules diffuse out of the cell and are washed away; the remaining green-fluorescent signal is a direct measure of the amount of active caspase that was present at the time the inhibitor was added. This kit includes the FLICA reagent specific for caspase-3 and -7, Hoechst 33342 stain, and propidium iodide stain, which allows the simultaneous evaluation of caspase activation, membrane permeability, and cell cycle by flow cytometry.

View a selection guide for all apoptosis assays for flow cytometry.

Dead Cell Apoptosis Kit with Annexin V Alexa Fluor™ 488 & Propidium Iodide (PI) (Invitrogen™)

The Dead Cell Apoptosis Kit with Annexin V Alexa Fluor™ 488 & Propidium Iodide is a flow cytometry kit used to measure early apoptosis by detecting phosphatidyl serine expression and membrane permeability.

View a selection guide for all apoptosis assays for flow cytometry.

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, which leads to higher binding efficiency, resulting in highly accurate characterization of the apoptotic process.

Multi-parametric
Many publications require a minimum of two different ways to identify that cells are apoptotic. This multi-parametric kit detects phosphatidyl serine (PS) on the cytoplasmic surface of the cell membrane and membrane integrity using propidium iodide.

How it works
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.

APO BrdU (TUNEL) Kit (Invitrogen™)

The ApoTarget™ APO-BRDU Kit is a two-color staining method for labeling DNA breaks and total cellular DNA to detect apoptotic cells by flow cytometry.

View a selection guide for all apoptosis assays for flow cytometry.

Vybrant™ FAM Caspase-8 Assay Kit, for flow cytometry (Invitrogen™)

The Vybrant™ FAM Caspase-8 Assay Kit employs a novel approach to detect active caspases that is based on a fluorescent inhibitor of caspases (FLICA™) methodology, essentially an affinity label. The reagent associates a fluoromethyl ketone (FMK) moiety, which can react covalently with a cysteine, with a caspase-specific amine acid sequence. For caspase-8, this recognition sequence is leucine-glutamic acid-threonine-aspartic acid (LETD). A fluorescein group is attached as a reporter. The FLICA reagent is thought to interact with the enzymatic reactive center of an activated caspase via the recognition sequence, and then to attach covalently through the FMK moiety. The FLICA inhibitor is cell permeant and noncytotoxic. Unbound FLICA molecules diffuse out of the cell and are washed away; the remaining green-fluorescent signal is a direct measure of the amount of active caspase that was present at the time the inhibitor was added. This kit includes the FLICA reagent specific for caspase-8, Hoechst 33342 stain, and propidium iodide stain, which allows the simultaneous evaluation of caspase activation, membrane permeability, and cell cycle by flow cytometry.

View a selection guide for all apoptosis assays for flow cytometry.

eBioscience™ Annexin V Apoptosis Detection Set PE-Cyanine7 (Invitrogen™)

Annexins are a family of calcium-dependent phospholipid-binding proteins that preferentially bind phosphatidylserine (PS) in all mammalian species. 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™ 660 or eFluor™ 780. 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).

Due to the emission spectrum of PE-Cyanine7, the Annexin V Apoptosis Detection Set PE-Cyanine7 is not compatible with propidium iodide and 7-AAD. It is recommended to substitute a Fixable Viability Dye such as eFluor™ 660 or eFluor™ 780 in their place.

Not included:
Fixable Viability Dye eFluor™ 660 (cat. 65-0864)
Fixable Viability Dye eFluor™ 780 (cat. 65-0865)

Note: Fixable Viability Dye eFluor™ 450 is not recommended for use with Annexin V Apoptosis Detection Kits.

Reactivity/Species
Human, Mouse, Rat

Reported Application
Flow Cytometric Analysis

Tali™ Apoptosis Kit - Annexin V Alexa Fluor™ 488 & Propidium Iodide (Invitrogen™)

The Tali® Apoptosis Kit enables identification of apoptotic cells and discrimination of apoptotic from necrotic and live cells in a population. The kit stains apoptotic cells with green Annexin V – Alexa Fluor® 488, stains necrotic cells with both red propidium iodide and green Annexin V – Alexa Fluor® 488, and does not stain live cells.

The Tali®Apoptosis Kit contains:

Annexin V – Alexa Fluor® 488 conjugate

• Annexin Binding solution
• Propidium Iodide (PI) solution

Apoptosis is a carefully regulated process of cell death that occurs as a normal part of development. Inappropriately regulated apoptosis is implicated in disease states, such as Alzheimer’s disease and cancer. Apoptosis is distinguished from necrosis by characteristic morphological and biochemical changes, including compaction and fragmentation of the nuclear chromatin, shrinkage of the cytoplasm, and loss of membrane asymmetry.

Easily Discriminate Apoptotic from Necrotic Cells with Two-color Staining
In normal live cells, phosphatidyl serine (PS) is located on the cytoplasmic surface of the cell membrane. In apoptotic cells, however, PS is translocated from the inner to the outer surface of the plasma membrane, exposing it to the extracellular environment. The human anticoagulant Annexin V displays a high affinity for PS. Annexin V labeled with a fluorophore such as Alexa Fluor® 488 can be used to identify apoptotic cells by binding to the exposed PS.

The Tali® Apoptosis Kit includes a solution of propidium Iodide (PI). PI is a cell-impermeant fluorogenic DNA-binding dye used for years to identify necrotic cells. PI is impermeant to live cells, but easily enters dead cells where it binds to nucleic acids and becomes fluorescent.

After staining a cell population using the Tali® Apoptosis Assay Kit with Annexin V-Alexa Fluor® 488 and Propidium Iodide, apoptotic cells display green fluorescence, dead cells display red and green fluorescence and live cells show little or no fluorescence.

Apoptotic DNA Ladder Kit (Invitrogen™)

The ApoTarget™ Quick Apoptotic DNA Ladder Detection Kit provides a simple and rapid procedure for extraction of chromosomal DNA. The procedure prepares DNA for use in the methods that detect DNA fragmentation in apoptotic cells. Unlike other kits requiring 1 to 2 days to finish, this detection method requires only less than 90 minutes to prepare DNA in a single tube without the need for extraction or column steps. DNA fragmentation can be easily visualized by agarose gel electrophoresis. This procedure increases recovery of small fragmented DNA, thereby improving the sensitivity of the assay.

Apo-BrdU Apoptosis Detection Kit (Invitrogen™)

The APO-BRDU™ Kit is a 2-color staining method for labeling DNA breaks and total cellular DNA to detect apoptotic cells by flow cytometry. The kit contains the instructions and reagents required for measuring apoptosis in cells, including positive and negative control cells for assessing reagent performance; washing, reaction and rinsing buffers for processing individual steps in the assay; terminal deoxynucleotidyl transferase enzyme (TdT), bromodeoxyuridine triphosphate (Br-dUTP), and fluorescein labeled anti-BrdU antibody for labeling DNA breaks and propidium iodide/RNase A solution for counterstaining the total DNA.

One of the most easily measured features of apoptotic cells is the break-up of the genomic DNA by cellular nucleases. These DNA fragments can be extracted from apoptotic cells and result in the appearance of DNA laddering when the DNA is analyzed by agarose gel electrophoresis. The DNA of non-apoptotic cells that remains largely intact does not display this laddering on agarose gels during electrophoresis. The large number of DNA fragments appearing in apoptotic cells results in a multitude of 3'-hydroxyl termini in the DNA. This property can be used to identify apoptotic cells by labeling the 3'-hydroxyl ends with bromolated deoxyuridine triphosphate nucleotides (Br-dUTP). The enzyme terminal deoxynucleotidyl transferase (TdT) catalyzes a template independent addition of deoxyribonucleoside triphosphates to the 3'-hydroxyl ends of double- or single-stranded DNA with either blunt, recessed or overhanging ends. A substantial number of these sites are available in apoptotic cells providing the basis for the method utilized in the APO-BRDU™ Kit. Recent evidence has demonstrated that Br-dUTP is more readily incorporated into the genome of apoptotic cells than are the deoxynucleotide triphosphates complexed to larger ligands like fluorescein, biotin or digoxigenin. This greater incorporation gives rise to a brighter flow cytometry signal when the Br-dUTP sites are identified by a fluorescein labeled anti-BrdU monoclonal antibody. Non-apoptotic cells do not incorporate significant amounts of the Br-dUTP due to the lack of exposed 3'-hydroxyl DNA ends.

Sufficient reagents are provided to process a total of 60 cell suspensions including 5 mL positive and 5 mL negative control cell suspensions of approximately 1x106 cells per mL in 70% (v/v) ethanol.

Reported Application
Immunocytochemistry, Flow Cytometric Analysis

APO-BrdU™ TUNEL Assay Kit, with Alexa Fluor™ 488 Anti-BrdU (Invitrogen™)

In collaboration with Phoenix Flow Systems, we offer the APO™-BrdU TUNEL Assay Kit which provides all the materials necessary to label and detect the DNA strand breaks of apoptotic cells. When DNA strands are cleaved or nicked by nucleases, a large number of 3´hydroxyl ends are exposed. In the APO-BrdU assay, these ends are labeled with BrdUTP and terminal deoxynucleotidyl transferase (TdT) using the TUNEL technique. Once incorporated ino the DNA, BrdU is detected using a bright and photostable green-fluorescent Alexa Fluor™ 488 dye-labeled anti-BrdU antibody.

View a selection guide for all apoptosis assays for flow cytometry.

Click-iT™ Plus TUNEL Assay for In Situ Apoptosis Detection, Alexa Fluor™ 488 dye (Invitrogen™)

The Click-iT® Plus TUNEL assay detects apoptotic cells in tissue and cultured cell samples through the use of a small, highly specific labeling moiety and a bright fluorescent dye. After incorporation of the labeling moiety into DNA fragments, detection is achieved through a catalyzed “click” reaction using conditions mild enough to preserve the fluorescent signal from GFP or RFP.

More tools for apoptosis detection and measurement >

• Optimized for the detection of apoptotic cells in either tissue or cell samples
• Multiplex enabled—optimized to work with fluorescent dyes or proteins such as GFP and RFP
• Improved TUNEL assay—better label incorporation due to small reactive moiety
• Bright apoptotic signal—uses Alexa Fluor® dyes, resulting in a stable, non-photobleaching fluorescent signal
• Flexibility—the assay can be configured to test from 1 to 50 samples at a time

Fragmentation of cellular DNA is a hallmark of apoptosis. The TUNEL assay is the most widely used method to detect fragmented DNA in apoptotic cells in tissue samples. The TUNEL assay begins with incorporation of modified dUTP at the 3’-OH end of the fragmented DNA. The dUTP modification is often the addition of a fluorophore. And due to the size of the fluorophore, the modified dUTP can display lower than expected incorporation rates. These lower incorporation rates can affect the sensitivity of the TUNEL assay. Additionally, the fluorophores used in currently available TUNEL assays suffer from photobleaching and fluorescent spectral overlap issues, both of which reduce sensitivity and ability to multiplex the assay.

The Click-iT® Plus TUNEL assay was developed to address these issues. The assay uses dUTP modified with an alkyne group (a small bio-orthogonal functional group) that allows the nucleotide to be more readily incorporated. After incorporation, a highly specific click reaction between the alkyne group and an Alexa Fluor® picolyl azide fluorescent dye, and subsequent detection of that dye, results in a sensitive and specific assay for the detection of apoptotic cells. Because of the gentle reaction conditions, the Click-iT® Plus TUNEL assay enables multiplexing with fluorescent proteins or dyes.

The Click-iT® Plus TUNEL assay has been validated with several different formalin-fixed paraffin-embedded tissue types. In all cases the ability to multiplex with fluorescent proteins and dyes was preserved. Additionally, the ability to stain actin using fluorescent-labeled phalloidin was also preserved.

The Click-iT® Plus TUNEL assay contains all of the reagents needed to detect apoptotic cells from either tissue or cell samples. The reagents supplied in this kit can be used to test 50 samples and can be configured to test from 1 to 50 samples at a time.

MitoProbe™ TMRM Assay Kit for Flow Cytometry

The MitoProbe TMRM Assay Kit contains tetramethylrhodamine methyl ester (TMRM) for the detection of mitochondrial membrane potential state and CCCP for the induction of mitochondrial membrane depolarization, if desired.

• Study mitochondrial membrane potential in live cells
• Quick, easy protocol

Determining the state of mitochondrial health can be achieved by detecting changes in the mitochondrial membrane potential. In a healthy cell with active mitochondria, TMRM is readily sequestered, thereby emitting a red-orange fluorescent signal. When apoptosis is induced or a treatment like CCCP is applied, the mitochondrial membrane is depolarized and the TMRM signal diminishes.

Mitochondrial Membrane Potential Apoptosis Kit, with Mitotracker™ Red & Annexin V Alexa Fluor™ 488, for flow cytometry (Invitrogen™)

This flow cytometry product provides a rapid and convenient assay for two hallmarks of apoptosis - phosphatidylserine externalization and changes in mitochondrial membrane potential. After staining a cell population with Alexa Fluor™ 488 annexin V and MitoTracker™ Red CMXRos dye in the provided binding buffer, live cells exhibit very little green fluorescence and bright red fluorescence, whereas apoptotic cells exhibit green fluorescence and decreased red fluorescence. These populations can easily be distinguished using a flow cytometer, and the 488 nm line of an argon-ion laser can be used to excite both dyes.

View a selection guide for all apoptosis assays for flow cytometry.

ApoDETECT Annexin V-FITC Kit (Invitrogen™)

The ApoDETECT Annexin V-FITC Kit provides all the reagents needed to detect the apoptotic cells. Annexin V is a Ca2+-dependent phospholipid binding protein. This protein can bind to a variety of phospholipids, but it has the highest affinity for phosphatidylserine (PS). The ApoDETECT Annexin V-FITC Kit binds to negatively charged phospholipid surfaces in a Ca2+-dependent manner. It also prevents the formation of the prothrombinase complex, thereby inhibiting formation of thrombin (anticoagulant activity). The ApoDETECT Annexin V-FITC Kit can be used to detect apoptotic cells by flow cytometry or immunofluorescent cytology.

Kit Attributes:

Applications: Validated application for ApoDETECT Annexin V-FITC Kit is detecting apoptotic cells by flow cytometry or immunofluorescent cytology.
Format: Vial(s)
Detection Method: Fluorescence microscopy
Product Size: One kit

In normal cells phosphatidylserine (PS) is located on the inner leaflet of the plasma membrane. During the early stages of apoptosis, PS is translocated to the outer layer and is exposed on the external surface of the cell. This early event in apoptosis can be detected by using a sensitive method in which to detect PS exposure. Translocation of PS to the external surface of the plasma membrane is not a unique property of apoptotic cells, as this phenomenon also occurs during cell necrosis. However, during apoptosis, the cell membrane remains intact; whereas, during necrosis, the cell becomes leaky and loses its integrity. Therefore, it is necessary to assess membrane integrity together with PS translocation.

Annexin V is a Ca2+-dependent phospholipid binding protein. This protein can bind to a variety of phospholipids, but it has the highest affinity for phosphatidylserine (PS). Based on its affinity for PS, Annexin V can be utilized as a sensitive probe for cell surface exposure of PS. To use the Annexin V protein as a probe for apoptotic cells, the protein has been labeled with fluorescein isothiocyanate (FITC). In this form, the protein can be used directly for quantification of apoptotic cells. The measurement of Annexin V binding when performed simultaneously with a dye exclusion test (such as propidium iodide) can be used to effectively discriminate between apoptotic and necrotic cells.

EnzChek™ Caspase-3 Assay Kit #1, Z-DEVD-AMC substrate (Invitrogen™)

The EnzChek Caspase-3 Assay Kit #1 allows detection of apoptosis by providing a simple and reliable method for assaying caspase-3 activity. The basis for the assay is the aminomethylcoumarin (AMC)-derived substrate Z-DEVD-AMC. This substrate, which is weakly fluorescent in the UV range (excitation/emission maxima ~330/390 nm), yields a bright, blue-fluorescent product (excitation/emission maxima ~342/441 nm) upon proteolytic cleavage. The kit can be used to continuously measure the activity of caspase-3 and closely related proteases in cell extracts and purified enzyme preparations, using a fluorometer or fluorescence microplate reader.

Membrane Permeability Dead Cell Apoptosis Kit with PO-PRO™-1 and 7-Aminoactinomycin D, for flow cytometry (Invitrogen™)

This flow cytometry product detects apoptosis on the basis of changes that occur in the permeability of cell membranes using PO-PRO™-1 and 7-aminoactinomycin D (7-AAD) nucleic acid stains. After staining a cell population with PO-PRO™-1 dye and 7-AAD, apoptotic cells show violet fluorescence, dead cells show violet and red fluorescence, and live cells show little or no fluorescence. These populations can easily be distinguished by a flow cytometer that uses both a violet laser and the 488 nm line of an argon-ion laser for excitation. Each kit contains sufficient reagents for approximately 200 flow cytometry tests.

View a selection guide for all apoptosis assays for flow cytometry.