FlowCellect™ DNA Damage Histone H2A.X Dual Detection Kit

FlowCellect™ DNA Damage Histone H2A.X Dual Detection Kit

FlowCellect

25 Tests / Kit

Millipore’s FlowCellect™ Dual Detection kits are a series of flow cytometry products which include a pair of antibodies that bind to the same protein; one to detect total protein expression and another to detect the phosphorylated form of the same target. Using two parameter analysis, we can achieve target specific detection of phosphorylation and, by doing so, eliminate false positives while enhancing the signal to noise ratio.

FlowCellect™ Histone H2A.X DNA Damage Dual Detection kit includes two directly conjugated antibodies, a phospho-specific Anti-phospho-Histone H2A.X (Ser139)-PerCP and an Anti-Histone H2A.X-FITC conjugated antibody to measure total levels of Histone H2A.X. This two color kit is designed to detect the extent of Histone H2A.X pathway activation by measuring H2A.X phosphorylation relative to the total H2A.X expression in any given cell population. By doing such, the levels of both the total and phosphorylated protein can be measured simultaneously in the same cell, resulting in a normalized and accurate measurement of H2A.X activation after stimulation. Moreover, simultaneous measurement of both total and phospho-Histone H2A.X confirms target specificity of the phosphorylation event. Together, a total and phospho antibody duo performed in multiplex provides an enhanced and more reliable detection of the phospho: total ratio within a mixed cell population. Using this antibody pair provides a sensitive and valuable tool to study the factors that induce DNA damage and/or affect DNA repair, and allow one to explore the linkage between DNA damage, cell cycle checkpoints, and initiation of apoptosis.

H2A.X is a member of the histone H2A family. Histone H2A.X resides downstream of the DNA damage kinase signaling cascade. Phosphorylation of Histone H2A.X at serine 139 is an important indicator of DNA damage. As the level of DNA damage increases, the level of phospho Histone H2A.X (also known as γH2AX) increases, accumulating at the sites of DNA damage. This accumulation of phospho Histone H2A.X is often used to indicate the level of DNA damage present within the cell. H2AX is also responsible for recruiting response proteins to the site of DNA damage and may play a role in DNA repair.

In order to validate that the level of H2A.X activation is accurate, a total H2A.X antibody is multiplexed by flow cytometry to allow the total and phospho levels to be normalized since DNA content levels vary based on which stage cells are within the cell cycle. Histone doubles in content during the cell cycle at the same rate DNA content doubles. Having a total H2A.X will account for this increase, so the phosphorylated levels can be accurately determined.

In all, a comprehensive understanding of Histone H2A.X activity and understanding the consequence of its activation can provide the researcher with useful information which will be important in understanding the intrinsic nature of the DNA damage response. Millipore’s FlowCellect™ Histone H2A.X DNA Damage Dual Detection Kit is designed to allow the researcher to monitor and accurately measure phospho-specific Histone H2A.X activation in a population of cells.

Flow Cytometry

Human

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Lot to lot in-process testing on all kit components

• 1. 20X Anti-phospho-Histone H2A.X (Ser139)-PerCP: (Part No. CS206553) One vial containing 150 µL.
• 2. 20X Anti-Histone H2A.X-FITC: (Part No. CS206429) One vial containing 150 µL.
• 3. Fixation Buffer: (Part No. CS202122) One bottle containing 13 mL.
• 4. 10X Wash Buffer: (Part No. CS202123) One bottle containing 13 mL.
• 5. 5X Assay Buffer: (Part No. CS202124) One 55 mL bottle
• 6. 1X Permeabilization Buffer: (Part No. CS203284) One bottle containing 14 mL.

Mouse

4-8°C for antibodies and buffers

Fluorescent

FlowCellect™ DNA Damage Histone H2A.X Dual Detection Kit

Flow Cellect

ExpressPro & inCyte

Monoclonal Antibody

Research