TUNEL Assay Kit - BrdU-Red (ab66110)
Key features and details
- Assay type: Quantitative
- Detection method: Fluorescent
- Platform: Flow cytometer, Fluorescence microscope
- Assay time: 3 hr
- Sample type: Adherent cells, Suspension cells, Tissue
Overview
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Product name
TUNEL Assay Kit - BrdU-Red
See all DNA fragmentation kits -
Detection method
Fluorescent -
Sample type
Tissue, Adherent cells, Suspension cells -
Assay type
Quantitative -
Assay time
3h 00m -
Species reactivity
Reacts with: Mammals, Other species -
Product overview
TUNEL Assay Kit - BrdU-Red ab66110 uses a convenient and sensitive method to detect DNA fragmentation by flow cytometry and fluorescence microscopy in live cells.
The TUNEL assay is used to detect DNA fragmentation, such as in apoptosis. It uses terminal deoxynucleotidyl transferase (TdT) to catalyze the incorporation of deoxynucleotides at the free 3'-hydroxyl ends of fragmented DNA. The deoxynucleotides are then labeled in a variety of ways for detection of the degree of DNA fragmentation.
This TUNEL assay protocol is based on Br-dUTP (bromolated deoxyuridine triphosphate nucleotide), which can be more readily incorporated into DNA strand breaks by the TdT enzyme than other dUTP labels such as FITC, biotin or dioxigenin. The greater incorporation rate produces a brighter signal when the Br-dUTP sites are detected with an anti-BrdU monoclonal antibody directly labeled with a red fluorochrome.
The BrdU-Red signal can be analyzed at Ex/Em 488/576 nm, with an optional 7-AAD counterstain at Ex/Em 488/655nm.
This TUNEL assay kit includes both negative and positive control cells. It is designed to be suitable for studying DNA fragmentation in GFP-transfected cells.
Tunel assay protocol summary:
- fix cells / tissues with formaldehyde, or deparaffinize and rehydrate if paraffin sections, and wash
- incubate cells in 70% ethanol for 30 min at 4ºC, or if tissues incubate with proteinase K solution for 5 min at room temp and refix with formaldehyde
- wash
- incubate in DNA labeling solution for 60 min at 37ºC
- wash
- incubate in antibody solution for 30 min at room temp
- add 7-AAD / RNase A solution and incubate for 30 min at room temp
- analyze with flow cytometry or fluorescent microscopy -
Notes
This kit is BrdU-Red labeled (Ex/Em = 488/576 nm). It was previously called TUNEL Assay Kit - In situ BrdU-Red DNA Fragmentation.
To use FITC (Ex/Em = 495/519 nm) as a label, we recommend TUNEL Assay Kit - FITC (ab66108).
For chromogenic TUNEL staining, we recommend TUNEL Assay Kit - HRP-DAB ab206386.
Find out more about the TUNEL method in the TUNEL staining / TUNEL assay guide.
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Platform
Flow cytometer, Fluorescence microscope
Properties
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Storage instructions
Please refer to protocols. -
Components Identifier 60 tests 7-AAD/RNase Staining Buffer Amber bottle 1 x 30ml Anti-BrdU-Red Antibody Orange 1 x 300µl Br-dUTP Violet 1 x 480µl Negative Control Cells Neutral 1 x 5ml Positive Control Cells Brown 1 x 5ml Reaction Buffer Green 1 x 600µl Rinse Buffer Red 1 x 120ml TdT Enzymes 1 x 45µl Wash Buffer Blue 1 x 120ml -
Research areas
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Relevance
Internucleosomal DNA fragmentation is a hallmark of apoptosis in mammalian cells.
Images
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In Situ BrdU-Red DNA Fragmentation (TUNEL) Assay Kit (ab66110)
Detection of DNA fragmention (TUNEL staining) using the negative and positive control cells (HL-60 untreated and treated with camptothecin). Cells were stained following the assay protocol. The fluorescence signal was detected and analyzed using BD FACScan System (Becton Dickinson).
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Fluorescence Microscopy - In situ BrdU-Red DNA Fragmentation (TUNEL) Assay Kit (ab66110) This image is courtesy of an Abreview submitted by James GahanTUNEL staining in whole mount Hydractinia echinata using In situ BrdU-Red DNA Fragmentation (TUNEL) Assay Kit (ab66110).
Animals were fixed in 4% PFA in PBS for 1 hour and processed as per the protocol without proteinaseK treatment. In place of proteinaseK animals were permeabilised in 3% Triton in PBS for 15 minutes. Animals were counter-stained with DAPI.
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Immunohistochemistry - ab66110 Image from Hopkins J et al., PLoS Genet 10(7), fig 1C. Doi: 10.1371/journal.pgen.1004413. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/Immunohistochemical analysis of paraffin embedded 5 micron thick testis tissues of 8 week old Stag3+/− and Stag3−/− (Stromal antigen) mice . Apoptotic cells were detected using In situ BrdU-Red DNA Fragmentation (TUNEL) Assay Kit (ab66110). DAPI was used as a counterstain.
