Denmark Finland Norway Sweden

Product of the month - August

 

DNA Fragmentation

Apoptosis - Detection of DNA Fragmentation

Apoptosis is a natural biological phenomenon by which unwanted cells are eliminated during development and other physiological processes. In the last step of apoptosis, activated endonucleases (caspase-activated DNase) break down genomic DNA between nucleosomes, generating fragments of multiples of 180 base pairs in size (Figure 1). This biochemical hallmark of apoptosis is an irreversible event and commits the cell to die. In contrast, necrotic cell death is accompanied by late and random DNA fragmentation through the release of lysosomal DNases. Techniques that detect DNA fragmentation are thus not necessarily specific to apoptosis, but may detect DNA damage in a variety of cell death paradigms. Therefore they can be paired with additional assays if necessary.

From Roche Applied Science we offer several kits for the detection of DNA fragmentation. Both for single cell analysis and in cell populations.

Figure 1. Overview of the mechanism of DNA fragmentation
and the appearance of the “DNA ladder”.

Click on picture for larger view.

DNA Fragmentation in Individual Cells
The DNA breaks generated by the endonucleases can be detected by enzymatic labeling of the free 3’-OH termini. This end-labeling method, TUNEL (terminal dUTPnick end-labeling), enables highly sensitive detection of apoptosis in tissue and single cells.

The use of fluorescein-dUTP to label the DNA strand breaks allows the detection of the incorporated nucleotides directly with a fluorescence microscope or a flow cytometer. Furthermore, the fluorescence may be converted into a colorimetric signal with an anti-fluorescein antibody conjugated with a reporter enzyme (Figure 2).


 Click on picture for larger view

Detection method Product Size Cat.No. Price (SEK)
FACS/
Fluorescence microscopy		 In situ Cell Death Detection Kit, Fluorescein 50 tests  11684795910 

4 690
In situ Cell Death Detection Kit, TMR Red 50 tests 12156792910

4 690
Light microscopy In situ Cell Death Detection Kit, POD 50 tests 11684817910

5 175
In situ Cell Death Detection Kit, AP 50 tests 11684809910

5 175

DNA Fragmentation in Cell Populations

Studying apoptosis in cell populations can be done by detection of the histone-associated DNA fragments present in the cytoplasm of cells undergoing apoptosis. Using the Cell Death Detection ELISAPLUS (Figure 3.) necrotic and apoptotic cells can be distinguished by analyzing samples from the cell culture supernatant (DNA fragments from necrotic cells) and from a cell lysate (DNA fragments from apoptotic cells). The assay is highly sensitive (detects apoptosis in as few as 600 cells), does not require proliferating cells, and works on cells from a variety of species.

 


Click on picture for larger view

Product Size Cat.No. Price (SEK)
Cell Death Detection ELISAPLUS 96 tests 11774425001

4 200
10 x 96 tests 11920685001

23 455