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Comet SCGE assay kit

 
ADI-900-166 50 tests 237.00 USD
 
  • Ready-to-use Comet Slides allow direct application of sample without pretreatment
  • Shorter assay time allows for higher throughput sample analysis
  • Hydrophobic barrier allows sample treatment with DNA repair enzymes
  • Unique nucleic acid stain provides improved sensitivity for DNA visualization compared to ethidium bromide
Exposure of cells to oxidative and environmental stresses frequently results in the breakdown or oxidation of genomic DNA. Assays to evaluate the integrity of genomic DNA, or to assess the presence of oxidized DNA are frequently used as a means of verifying the onset of apoptosis or DNA damage. The COMET SCGE Assay measures DNA damage by fluorescently detecting the integrity of DNA liberated from cells embedded in low melting point agarose. Upon electrophoresis, fragmented DNA produces a characteristic "comet" shaped tail as small DNA fragments migrate in the gel more rapidly than in-tact genomic DNA. The COMET SCGE Assay is a fast and simple electrophoresis method to detect and quantitate DNA fragmentation in cells associated with DNA damage and apoptosis. A unique nucleic acid stain provides improved sensitivity for DNA visualization compared to ethidium bromide.

Product Specification

Assay Time:3 hours
 
Application:For the measurement of DNA Damage, DNA Repair, and Apoptosis in cells and tissue from any species via electrophoresis.
 
Use/Stability:Store all components ambient temperature.
 
Long Term Storage:Ambient
 
Kit/Set Contains:Lysis solution, Comet LMAgarose, Comet slide, EDTA
 
Miscellaneous/General:The COMET SCGE Assay is a single cell gel electrophoresis assay that provides a simple and effective method for evaluating DNA damage in cells. The principle of the assay is based upon the ability of denatured, cleaved DNA fragments to migrate out of the cell under the influence of an electric field, whereas undamaged DNA migrates slower and remains within the confines of the nucleoid when a current is applied. Evaluation of the DNA “comet” tail shape and migration pattern allows for assessment of DNA damage. In this assay, cells are immobilized in a bed of low melting point agarose on a Comet Slide. Following a gentle cell lysis, samples are treated with alkali to unwind and denature the DNA and hydrolyze sites of damage. The samples are then submitted to electrophoresis and staining with a fluorescent DNA intercalating dye. The sample is then visualized by epifluorescence microscopy. As an alternative for researchers who do not have access to a fluorescence microscope, silver staining allows standard light microscopy for data analysis. The electrophoresis step may be performed using TBE buffer or alkaline electrophoresis solution. TBE circumvents the problems associated with use of non-buffered alkali for electrophoresis, such as poor DNA migration due to saturation of charge by sodium ions, and difficulties in controlling voltage. TBE is preferred for analysis of apoptosis and enables use of the tail length, rather than the tail moment, for data analysis. The alkaline electrophoresis is more sensitive and will detect smaller amounts of damage. The electrophoresis time may be extended up to 40 minutes when running at very low amperage. Data may be analyzed qualitatively if the comets are scored according to categories of small to large tail lengths. Quantitative and statistical data can readily be generated by analysis of the results using one of several commercially available image analysis software packages which calculate tail length and tail moment. The COMET SCGE Assay’s Comet Slide is specially treated to promote adherence of low melting point agarose. This eliminates the time consuming and unreliable traditional method of preparing base layers of agarose. The use of Comet Slides shortens assay time, and allows the rapid and reliable analysis of large numbers of samples. SYBR® Green 1 provides improved sensitivity compared to ethidium bromide for DNA visualization.
 
Background / Technical Information:Please note that the following kit component, SYBR® Green 1 Nucleic Acid Gen Stain, is no longer offered as a kit component with ADI-900-166, COMET SCGE assay kit. Instead we recommended the kit component, ENZ-51014 -- Nuclear-ID® Green cell cycle kit for flow cytometry or ENZ-52401 -- Hoechst 33342 (ultra pure), which is not supplied with ADI-900-166.
 

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