TUNEL Assay Kit - Edu-Orange (ab252888)
Key features and details
- Detection method: Flow cytometry-fluorescent
- Platform: Flow cytometer, Fluorescence microscope
- Sample type: Adherent cells, Suspension cells, Tissue
Overview
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Product name
TUNEL Assay Kit - Edu-Orange
See all DNA fragmentation kits -
Detection method
Flow cytometry-fluorescent -
Sample type
Tissue, Adherent cells, Suspension cells -
Assay duration
Multiple steps standard assay -
Product overview
TUNEL Assay Kit - Edu-Orange ab252888 uses an EdU click chemistry method to detect fragemented DNA. The kit contains sufficient reagents to detect total/fragmented DNA in apoptotic cells in a 1 X 96-well plate or on 50 cover slips.
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 uses modified EdUTP nucleotides. A click reaction is then used to attach an orange dye (Ex 490 / Em 580, FL2 channel) to the EdUTP for detection by either flow cytometry or fluoresence microscopy. A green DNA staining dye is used for contrast (Ex 440 / Em 540)
TUNEL assay protocol summary:
- fix cells for 15 mins at room temp
- wash cells
- incubate with permeabilization buffer for 10 mins and wash twice
- add TUNEL buffer and incubate for 10 mins
- spin down cells and remove buffer
- add TUNEL reaction cocktail and incubate for 1hr at 37ºC
- spin down cells and remove cocktail, and wash
- add Click reaction cocktail, incubate for 30 mins and wash 3 times
- add DNA stain, incubate for 20 mins and wash
- analyze with flow cytometer or fluoresence microscope -
Tested applications
Suitable for: Flow Cyt, FMmore details -
Platform
Flow cytometer, Fluorescence microscope
Properties
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Storage instructions
Please refer to protocols. -
Components 100 tests Copper Reagent (100X) 1 x 100µl EdUTP DNA Label (50X) 1 x 100µl Fixative Solution 1 x 10ml Fluorescent Azide (100X) 1 x 100µl Permeabilization Buffer (10X) 1 x 25ml Reducing Agent (20X) 1 x 500µl Total DNA Stain (1000X) 1 x 10µl TUNEL Enzyme 1 vial TUNEL Enzyme Buffer 1 x 500µl TUNEL Reaction Buffer (10X) 1 x 1ml Wash Buffer (10X) 1 x 25ml -
Research areas
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Relevance
Internucleosomal DNA fragmentation is a hallmark of apoptosis in mammalian cells.
Images
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Example Data
Detection of TUNEL-positive apoptotic strand breaks. Jurkat (Human T cell leukemia cell line from peripheral blood) cells-DNAse treated (106 cells/ml) induced strand breaks.
Unstained cells w/vehicle (white), background control cells processed for click reaction (green), negative control (untreated cells, TUNEL and click reaction; blue), DNase-treated cells (pink).
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Example dataLeft panel: DNase treated HeLa (Human epithelial cell line from cervix adenocarcinoma) cells (105 cells/ ml). DNA staining and TUNEL and click reactions were performed.
Green: nuclear stain; Red (TUNEL positive); orange: apoptotic cells.
Right panel: DNase treated Jurkat (Human T cell leukemia cell line from peripheral blood) cells (106 cells/ ml). DNA staining and TUNEL and click reactions were performed.
Green: nuclear stain; Red (TUNEL positive); orange: apoptotic cells.