Invitrogen™ Apo-Direct Apoptosis Detection Kit
On demandInvitrogen™ Apo-Direct Apoptosis Detection Kit
The
APO-DIRECT™ 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), Fluorescein-deoxyuridine triphosphate 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 directly
conjugated fluorescein- deoxyuridine triphosphate nucleotides
(FITC-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-DIRECT™ Kit. Non-apoptotic cells do not
incorporate significant amounts of the FITC-dUTP due to the lack of
exposed 3'-hydroxyl DNA ends.
Sufficientreagents
are provided to process 50 cell suspensions including 5 mL positive and
5 mL negative control cell suspensions of approximately 1x106 cells per mL in 70% (v/v) ethanol.
There are no specifications
There are no report
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