ab23996 gave a positive result in the following lysates:
MRC5 whole cell lysate (SV40 transformed immortal (WT) fibroblasts), Hela whole cell, NIH3T3 whole cell, HepG2 whole cell and Jurkat whole cell.
Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
1/1000 - 1/5000. Detects a band of approximately 95 kDa (predicted molecular weight: 85 kDa). Abcam recommends using milk as the blocking agent. Abcam welcomes customer feedback and would appreciate any comments regarding this product and the data presented above.
Use a concentration of 1 µg/ml.
FunctionComponent of the MRE11-RAD50-NBN (MRN complex) which plays a critical role in the cellular response to DNA damage and the maintenance of chromosome integrity. The complex is involved in double-strand break (DSB) repair, DNA recombination, maintenance of telomere integrity, cell cycle checkpoint control and meiosis. The complex possesses single-strand endonuclease activity and double-strand-specific 3'-5' exonuclease activity, which are provided by MRE11A. RAD50 may be required to bind DNA ends and hold them in close proximity. NBN modulate the DNA damage signal sensing by recruiting PI3/PI4-kinase family members ATM, ATR, and probably DNA-PKcs to the DNA damage sites and activating their functions. It can also recruit MRE11 and RAD50 to the proximity of DSBs by an interaction with the histone H2AX. NBN also functions in telomere length maintenance by generating the 3' overhang which serves as a primer for telomerase dependent telomere elongation. NBN is a major player in the control of intra-S-phase checkpoint and there is some evidence that NBN is involved in G1 and G2 checkpoints. The roles of NBS1/MRN encompass DNA damage sensor, signal transducer, and effector, which enable cells to maintain DNA integrity and genomic stability. Forms a complex with RBBP8 to link DNA double-strand break sensing to resection. Enhances AKT1 phosphorylation possibly by association with the mTORC2 complex.
Tissue specificityUbiquitous. Expressed at high levels in testis.
Involvement in diseaseNijmegen breakage syndrome Breast cancer Aplastic anemia Defects in NBN might play a role in the pathogenesis of childhood acute lymphoblastic leukemia (ALL).
DomainThe FHA and BRCT domains are likely to have a crucial role for both binding to histone H2AFX and for relocalization of MRE11/RAD50 complex to the vicinity of DNA damage. The C-terminal domain contains a MRE11-binding site, and this interaction is required for the nuclear localization of the MRN complex. The EEXXXDDL motif at the C-terminus is required for the interaction with ATM and its recruitment to sites of DNA damage and promote the phosphorylation of ATM substrates, leading to the events of DNA damage response.
Post-translational modificationsPhosphorylated by ATM in response of ionizing radiation, and such phosphorylation is responsible intra-S phase checkpoint control and telomere maintenance.
Cellular localizationNucleus. Nucleus, PML body. Chromosome, telomere. Localizes to discrete nuclear foci after treatment with genotoxic agents.
ICC/IF image of ab23996 stained HeLa cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab23996, 1µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 100% methanol fixed (5 min) HepG2 and MCF7 cells at 1µg/ml.