Product nameChromatin Condensation Assay Kit
Sample typeAdherent cells, Suspension cells
Species reactivityReacts with: Other species, Mammals
The Chromatin Condensation Assay Kit (ab139479) provides a convenient approach for analysis of late stage apoptosis by flow cytometry and microscopy, and it can also be used in microplate assays. The basis of the assay is that the compacted chromatin of apoptotic cells binds higher amounts of nuclear dye compared to the healthy cells. The kit is suitable for differentiating between healthy and apoptotic cells with condensed nuclei.
A control apoptosis inducing agent, Staurosporine, is provided for monitoring apoptotic changes in nuclear organization. Potential applications for live-cell studies include quickly testing cell cultures for overall health, as well as rapid screening for compounds that induce apoptosis and chromatin condensation.
The cell-permeable dye used in this application is an aromatic, planar cationic structure that inserts between stacked base pairs on the DNA duplex, providing an environmentally-dependent fluorescence enhancement of the dye molecules and large increases in fluorescence signal relative to the free dye in solution. Since the signal enhancement provides a proportional response, direct quantitation of DNA is possible. Further signal increase is observed upon DNA condensation during apoptosis. Considering the general mutagenic effect of nucleic acid-binding dyes, careful storage and handling of this dye is recommended.
PlatformMicroplate reader, Fluor. microscope, Flow cyt.
Storage instructionsPlease refer to protocols.
Components 200 tests 10X Assay Buffer 1 x 30ml Apoptosis inducer (Staurosporine) 1 vial Green Cell Cycle Detection Reagent 1 x 100µl
- FCCP, mitochondrial oxidative phosphorylation uncoupler (ab120081)
- Cycloheximide, Protein synthesis inhibitor (ab120093)
- SB431542, ALK inhibitor (ab120163)
- Z-VAD(OH)-FMK, Irreversible general caspase inhibitor (ab120382)
- Z-VAD(OMe)-FMK, Cell permeable, irreversible pan-caspase inhibitor (ab120487)
- Dorsomorphin (Compound C), AMP-kinase inhibitor (ab120843)
- Calcein AM, fluorescent dye for cell viability (ab141420)
Raw data is presented in panels A and B where suggested gating is indicated by rectangles drawn around the cell populations. Panels C and D represent the separation of healthy and apoptotic nuclei based on their characteristic fluorescence.
Detection of the Sub-G0 phase in permeabilized cells using Green Cell Cycle Detection dye. In the permeabilized cells, the dye intercalates with the chromosomal DNA, resulting in a highly fluorescent healthy cell population. Upon Staurosporine treatment, a population with lower fluorescence, corresponding to fragmented DNA, can be detected via flow cytometry.
Absorption and fluorescence emission spectra for Green Cell Cycle Detection dye. Spectra were determined in 1X Assay Buffer.
ab139479 has not yet been referenced specifically in any publications.