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Phospho-Histone H2A.X (Ser139) Antibody (12-9865-42) in Flow
Normal human peripheral blood cells were stimulated with Anti-CD3 and Anti-CD28 Functional Grade Purifieds (Product # 16-0037 and 16-0289) for 3 days. Cells were then unstimulated (left) or stimulated with etoposide (right) for 4 hours and then intracellularly stained with Anti-Human PARP1 (Cleaved) Alexa Fluor® 488 (Product # 53-6668-42) and Anti-Human/Mouse phospho-H2AX (S139) PE using the Foxp3/Transcrip... View More
Please note: We are reviewing Western blot images included in the antibody testing data in our catalog, including those provided by third parties. Unless expressly labeled or annotated as “raw-unedited”, Western blot images included in the antibody testing data in our catalog may have been edited, optimized or otherwise adjusted for presentation.
Product Details
12-9865-42
Applications
Tested Dilution
Publications
Product Specifications
Species Reactivity
Human,
Mouse
Published species
Human,
Mouse
Host/Isotype
Mouse
/ IgG1, kappa
Recommended Isotype Control
Class
Monoclonal
Type
Antibody
Clone
CR55T33
Conjugate
PE
PE
PE
Excitation/Emission Max
565/576 nm
View spectra
Form
Liquid
Concentration
5 µL/Test
Purification
Affinity chromatography
Storage buffer
PBS, pH 7.2, with BSA
Contains
0.09% sodium azide
Storage conditions
4°C, store in dark, DO NOT FREEZE!
Shipping conditions
Ambient (domestic); Wet ice (international)
RRID
Product Specific Information
Description: The CR55T33 monoclonal antibody recognizes phosphorylated serine 139 of human and mouse H2AX. H2AX is a member of the H2A histone family that complex with DNA and other histones to form the repeating nucleosome units characteristic of eukaryotic chromatin. Nucleosomes consist of approximately 147 base pairs of DNA wrapped around an octamer of histones composed of two each of the four histone proteins: H2A, H2B, H3 and H4. After induction of DNA damage such as double-strand breaks by irradiation, genotoxic stresses, replication errors or gene recombination, PI3K-like kinases (e.g., ataxia telangiectasia mutated (ATM), ataxia telangiectasia Rad-3-related (ATR), and DNA-dependent protein kinase (DNA-PK) are activated to phosphorylate serine 139 in H2AX. This early phosphorylation event plays a critical role in recruiting proteins involved in DNA repair.
The monoclonal antibody CR55T33 recognizes a single band of approximately 15 kDa on reduced cell lysates from Jurkat cells stimulated with etoposide.
Applications Reported: This CR55T33 antibody has been reported for use in intracellular staining followed by flow cytometric analysis.
Applications Tested: This CR55T33 antibody has been pre-titrated and tested by intracellular staining and flow cytometric analysis of stimulated normal human peripheral blood cells using the Foxp3/Transcription Factor Staining Buffer Set (Product # 00-5523-00) and protocol. Please refer to BestProtocols®: Protocol B: One step protocol for (nuclear) intracellular proteins located under the Resources Tab online. This can be used at 5 µL (0.25 µg) per test. A test is defined as the amount (µg) of antibody that will stain a cell sample in a final volume of 100 µL. Cell number should be determined empirically but can range from 10^5 to 10^8 cells/test.
Excitation: 488-561 nm; Emission: 578 nm; Laser: Blue Laser, Green Laser, Yellow-Green Laser.
Filtration: 0.2 µm post-manufacturing filtered.
Target Information
Histone H2A.X (H2AX) is a member of the histone H2A family which is one of the four core histones making up the nucleosome core particle. In eukaryotes, DNA double strand breaks (DSBs) have been shown to trigger the phosphorylation of serine 139 at the carboxy terminus of histone H2AX resulting in gamma-H2AX. The phosphorylation of H2AX can be detected by Western blotting or immunofluorescence, revealing the frequency of DSBs. The phosphatidylinositol 3-kinases have been implicated in H2AX phosphorylation, but it is unclear if ATM is the primary H2AX kinase or if other members of the family such as DNA-PK and ATR contribute in a similar manner. Structurally, H2A.x contains 143 amino acid residues. Histone H2A.X is considered a basal histone, being synthesized in G1 as well as in S-phase, and its mRNA contains polyA addition motifs and a polyA tail along with the conserved stem-loop and U7 binding sequences involved in the processing and stability of replication type histone mRNAs. There are two forms of Histone H2A.X mRNA, one about 1600 bases long and contains polyA; the other about 575 bases long, lacking polyA. The short form behaves as a replication type histone mRNA, while the longer behaves as a basal type histone mRNA. Histone H2A.X maps to the 11q23.2-q23.3 region of the human chromosome. Histone H2A.x contributes to histone-formation and therefore the structure of DNA. Histone H2A variant H2A.x specifically regulates the interaction of MDC1 (mediator of DNA damage checkpoint protein 1), a DNA repair protein to the sites of DNA damage.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
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Bioinformatics
Protein Aliases: H2A histone family member X; H2A histone family, member X; H2a/x; H2AFX; H2AX histone; Histone 2A.X; histone 5 protein 2ax; histone H2A.x; Histone H2a/x; unnamed protein product
Gene Aliases: gammaH2ax; H2A.X; H2A/X; H2AFX; Hist5-2ax
UniProt ID: (Human) P16104, (Mouse) P27661
Entrez Gene ID: (Human) 3014, (Mouse) 15270
DNA damage checkpoint
double-strand break repair via homologous recombination
DNA repair
cellular response to DNA damage stimulus
spermatogenesis
heterochromatin assembly
protein K63-linked ubiquitination
cellular response to gamma radiation
positive regulation of double-strand break repair via homologous recombination
protein localization to site of double-strand break
double-strand break repair
nucleosome assembly
response to ionizing radiation
positive regulation of DNA repair
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