Nexcelom Bioscience
Cellometer Cell-Based Assay Kits for the Vision CBA Analysis System
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Cellometer Annexin V-FITC / PI Apoptosis Reagents

Cellometer Annexin V-FITC / PI Apoptosis Reagents
Apoptosis, or programmed cell death, is a natural process of cellular self-destruction. Apoptosis is necessary for routine cell turnover and tissue homeostasis. It is also important in embryogenesis, maintenance of immune tolerance, and development of the nervous system. Problems with the regulation of apoptosis are thought to be linked to many cancers, degenerative diseases, and autoimmune diseases, making apoptosis a key target in many fields of clinical research.

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Figure 1

Scatter plots following Cellometer Vision CBA Analysis
Figure 1. Scatter plots following Cellometer Vision CBA Analysis show an increase in the apoptotic cell population from 4% to 22% following incubation with 10µm α-Tos, an apoptosis-inducing drug.

Principle of the Annexin V-FITC / PI Apoptosis Assay

Annexin V is a member of the annexin family of intracellular proteins that binds to phosphatidylserine (PS). PS is normally only found on the intracellular leaflet of the plasma membrane in healthy cells, but during apoptosis, PS translocates to the external leaflet. Fluorochrome-labeled Annexin V can then be used to specifically target and bind the PS on the surface of apoptotic cells. Cells undergoing necrosis following tissue damage or disease have compromised membranes that will also allow binding of PS. An additional dye is utilized to help differentiate necrotic cells within the population.

Propidium Iodide (PI) is a membrane-exclusion dye that permeates cells with compromised cell membranes and binds to DNA. Early apoptotic and healthy cells with intact membranes will exclude PI, while very late stage apoptotic and necrotic cells with compromised membranes are stained. Use of both Annexin V-FITC and PI allows researchers to characterize a cell population based on % normal, % apoptotic, and % necrotic /very late-stage apoptotic cells.

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Cellometer Calcein-AM / PI Vitality / Viability Kit

Cellometer Calcein-AM / PI Vitality / Viability Kit
The Cellometer Calcein-AM/PI Cell Vitality and Viability kit enables measurement of the number and concentration of both metabolically active and dead, or non-viable, cells in a population. The software automatically calculates percent vitality/viability based on the number of Calcein-positive (metabolically active) cells, and the number of PI-positive (non-viable) cells.

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Assay Principle

Acetoxymethyl ester of calcein (calcein-AM) is a non-fluorescent dye that passively enters cells. In metabolically active cells, the dye is converted by cytosolic esterases into green fluorescent calcein. The fluorescent calcein is retained by cells with intact membranes. Metabolically-active cells fluoresce in the green channel. Propidium iodide (PI) is a DNA-binding dye that enters cells with compromised membranes. Dead (compromised) nucleated cells fluoresce in the red channel.

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Cellometer Caspase-3 Apoptosis Kit

Cellometer Caspase-3 Apoptosis Kit
Caspase-3 belongs to a family of evolutionally conserved cysteine proteases that play a key role in regulating programmed cell death, or apoptosis. Caspase-3 activation triggers a series of irreversible events that lead to the death of the cell, including activation of the CAD endonuclease that degrades DNA within the nucleus and initiates chromatin condensation. Increasing evidence suggests involvement of caspase-3 in Alzheimer's Disease (AD) via cleavage of amyloid prescursor protein (APP) and tau and the formation of beta-amyloid (Aβ) and neurofibrillary tangles (NFTs). Other studies have demonstrated a role for caspase-3 in growth stimulation via PGE2. High amounts of caspase-3 have been linked to an increased rate of cancer recurrence and death in patients undergoing radiation treatment through the stimulation of growth of surviving cancer cells, making caspase-3 a key prognostic marker and therapeutic target in cancer therapy.

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Figure 2

Caspase-3 histogram

Table 1


Cell Population
% of Cells
Concentration (10∧6 cells/mL)
Total
100
2.99
Caspase +
40.16
1.20
Caspase -
59.84
1.79
Figure 2 and Table 1. Caspase-3 Activity histogram and data table for a Jurkat cell sample following overnight incubation with 10µm α-Tos, an apoptosis-inducing drug.

Principle of the Cellometer Caspase-3 Apoptosis Assay

The CaspGLOW™Fluorescein Active Caspase-3 Staining Kit offers a simple, sensitive method for detection of active Caspase-3 in living cells. When detecting caspase-3, we utilize a specific FITC-conjugated caspase-3 inhibitor, DEVD-FMK. DEVD-FMK is cell-permeable and binds irreversibly to active caspase-3 within the apoptotic cell. The fluorescent marker (FITC) on the FITC-DEVD-FMK allows for the detection of cells undergoing apoptosis via Caspase-3 activation using image cytometry. Each kit contains ready-to-use reagents sufficient for 100 tests along with a detailed instruction manual outlining cell staining, imaging, and cell cycle data analysis using the Cellometer Vision CBA Analysis System.

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Cellometer Caspase-8 Apoptosis Kit

Cellometer Caspase-8 Apoptosis Kit
The Cellometer Caspase-8 Apoptosis Kit is designed for fast, simple determination of Caspase-8 activity for a cell population using image cytometry. Each kit contains ready-to-use reagents sufficient for 100 tests along with a detailed instruction manual outlining cell staining, imaging, and cell cycle data analysis using the Cellometer Vision CBA Analysis System.

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Figure 3

Caspase-8 histogram

Table 2


Cell Population
% of Cells
Concentration (10∧6 cells/mL)
Total
100
3.88
Caspase +
27.54
1.07
Caspase -
72.17
2.80
Figure 3 and Table 2. Caspase-8 Activity histogram and data table for a Jurkat cell sample

Principle of the Cellometer Caspase-8 Apoptosis Assay

The CaspGLOW™Fluorescein Active Caspase-8 Staining Kit offers a simple, sensitive method for detection of active Caspase-8 in living cells. When detecting caspase-8, we utilize a specific FITC-conjugated caspase-8 inhibitor, IETD-FMK. IETD-FMK is cell-permeable and binds irreversibly to active caspase-8 within the cell. The fluorescent marker (FITC) on the IETD-FMK allows for the detection of cells undergoing apoptosis via Caspase-8 activation using the Vision CBA Analysis System. Results are presented as a Caspase positive population percentage and cell concentration.

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Cellometer PI Cell Cycle Kit

Cellometer PI Cell Cycle Kit
The Cellometer PI Cell Cycle Kit is designed for fast, simple determination of the cell cycle distribution for a cell population using image cytometry. The kit is specifically developed for use with the Cellometer Vision CBA Analysis System. Each kit contains ready-to-use PI/RNase staining solution sufficient for 100 tests along with a detailed instruction manual outlining cell staining, imaging, and cell cycle data analysis.

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Figure 4

Cell cycle histogram

Table 3


Cell Population
% of Gated Cells
CV
Concentration (10∧6 cells/mL)
Total
100
9.2
Sub G1
1.2
5.1
0.1
G0/G1
63.4
10.5
5.8
S
13.9
11.3
1.3
G2/M
21.0
8.2
1.9
Figure 4 and Table 3. Cell cycle histogram and data table for a Jurkat cell control sample

Principle of the Cellometer PI Cell Cycle Assay

Cell cycle analysis is used to determine the proportion of cells in each stage of the cell cycle for a given cell population based on variations in DNA content. In this assay, each cell is stained with propidium iodide (PI), a fluorescent nuclear staining dye that intercalates with DNA. Because the dye cannot enter live cells, the cells are fixed with ethanol or methanol, then all of the cells are stained. Cells preparing for division will contain increasing amounts of DNA and display proportionally increased fluorescence. Differences in fluorescence intensity are used to determine the percentage of cells in each phase of the cell cycle.
Cell cycle analysis

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Ordering Information

Catalog # Description Instrument Compatibility Size
Unit
Instruction Booklet
MSDS
U.S. Price
Cellometer Annexin V-FITC / PI Apoptosis Reagents  
CS1-0114 BioLegend Annexin V-FITC Cellometer Vision CBA 100 tests
each
 
--
$198.00
CS0-0115-100ML BioLegend Annexin V Binding Buffer Cellometer Vision CBA ~500 tests
each
 
--
$36.00
CS1-0116 Cellometer PI Staining Solution Cellometer Vision CBA 100 tests
each
 
$90.00
Cellometer Calcein-AM / PI Vitality / Viability Kit
CSK-0118 Calcein-AM/PI Cell Vitality/Viability Kit Vision, Vision CBA, Auto 2000 100 tests
each
$287.00
Cellometer Caspase-3 Apoptosis Kit
K183-100-N CaspGLOW™Fluorescein Active Caspase-3 Staining Kit Cellometer Vision CBA 100 tests
each
 
--
$404.00
K183-25-N CaspGLOW™Fluorescein Active Caspase-3 Staining Kit Cellometer Vision CBA 25 tests
each
 
--
$143.00
Cellometer Caspase-8 Apoptosis Kit
K188-100-N CaspGLOW™Fluorescein Active Caspase-8 Staining Kit Cellometer Vision CBA 100 tests
each
 
--
$404.00
K188-25-N CaspGLOW™Fluorescein Active Caspase-8 Staining Kit Cellometer Vision CBA 25 tests
each
 
--
$143.00
Cellometer PI Cell Cycle Kit
CSK-0112 Cellometer PI Cell Cycle Kit Cellometer Vision CBA 100 tests
each
$129.00

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