产品名称Anti-APE1抗体[13B8E5C2] - ChIP Grade
参阅全部 APE1 一抗
描述小鼠单克隆抗体[13B8E5C2] to APE1 - ChIP Grade
特异性This antibody is specific to the human APE/ref-1 protein.
经测试应用适用于: ICC/IF, IHC-Fr, IP, WB, ELISA, ICC, IHC-P, ChIPmore details
种属反应性与反应: Mouse, Rat, Human
Full length native protein (purified) (Human).
- WB: HepG2, HEK293 and wild-type HAP1 cell lysates. IHC-P: Human normal placenta tissue.
常规说明APE appears to form a unique link between the DNA base excision pathway, oxidative signalling, trancription regulation, cancer and cell-cycle control.
存放说明Shipped 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.
存储溶液Preservative: 0.05% Sodium azide
纯度Tissue culture supernatant
ChIP Related Products
Our Abpromise guarantee covers the use of ab194 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ICC/IF||Use at an assay dependent concentration.|
|IP||Use at an assay dependent concentration.|
|WB||1/2000. Predicted molecular weight: 35.6 kDa.|
|ICC||Use at an assay dependent concentration.|
|ChIP||Use at an assay dependent concentration.|
功能Multifunctional protein that plays a central role in the cellular response to oxidative stress. The two major activities of APEX1 in DNA repair and redox regulation of transcriptional factors. Functions as a apurinic/apyrimidinic (AP) endodeoxyribonuclease in the DNA base excision repair (BER) pathway of DNA lesions induced by oxidative and alkylating agents. Initiates repair of AP sites in DNA by catalyzing hydrolytic incision of the phosphodiester backbone immediately adjacent to the damage, generating a single-strand break with 5'-deoxyribose phosphate and 3'-hydroxyl ends. Does also incise at AP sites in the DNA strand of DNA/RNA hybrids, single-stranded DNA regions of R-loop structures, and single-stranded RNA molecules. Has a 3'-5' exoribonuclease activity on mismatched deoxyribonucleotides at the 3' termini of nicked or gapped DNA molecules during short-patch BER. Possesses a DNA 3' phosphodiesterase activity capable of removing lesions (such as phosphoglycolate) blocking the 3' side of DNA strand breaks. May also play a role in the epigenetic regulation of gene expression by participating in DNA demethylation. Acts as a loading factor for POLB onto non-incised AP sites in DNA and stimulates the 5'-terminal deoxyribose 5'-phosphate (dRp) excision activity of POLB. Plays a role in the protection from granzymes-mediated cellular repair leading to cell death. Also involved in the DNA cleavage step of class switch recombination (CSR). On the other hand, APEX1 also exerts reversible nuclear redox activity to regulate DNA binding affinity and transcriptional activity of transcriptional factors by controlling the redox status of their DNA-binding domain, such as the FOS/JUN AP-1 complex after exposure to IR. Involved in calcium-dependent down-regulation of parathyroid hormone (PTH) expression by binding to negative calcium response elements (nCaREs). Together with HNRNPL or the dimer XRCC5/XRCC6, associates with nCaRE, acting as an activator of transcriptional repression. Stimulates the YBX1-mediated MDR1 promoter activity, when acetylated at Lys-6 and Lys-7, leading to drug resistance. Acts also as an endoribonuclease involved in the control of single-stranded RNA metabolism. Plays a role in regulating MYC mRNA turnover by preferentially cleaving in between UA and CA dinucleotides of the MYC coding region determinant (CRD). In association with NMD1, plays a role in the rRNA quality control process during cell cycle progression. Associates, together with YBX1, on the MDR1 promoter. Together with NPM1, associates with rRNA. Binds DNA and RNA.
序列相似性Belongs to the DNA repair enzymes AP/ExoA family.
结构域The N-terminus contains the redox activity while the C-terminus exerts the DNA AP-endodeoxyribonuclease activity; both function are independent in their actions. An unconventional mitochondrial targeting sequence (MTS) is harbored within the C-terminus, that appears to be masked by the N-terminal sequence containing the nuclear localization signal (NLS), that probably blocks the interaction between the MTS and Tom proteins.
翻译后修饰Phosphorylated. Phosphorylation by kinase PKC or casein kinase CK2 results in enhanced redox activity that stimulates binding of the FOS/JUN AP-1 complex to its cognate binding site. AP-endodeoxyribonuclease activity is not affected by CK2-mediated phosphorylation. Phosphorylation of Thr-233 by CDK5 reduces AP-endodeoxyribonuclease activity resulting in accumulation of DNA damage and contributing to neuronal death.
Acetylated on Lys-6 and Lys-7. Acetylation is increased by the transcriptional coactivator EP300 acetyltransferase, genotoxic agents like H(2)O(2) and methyl methanesulfonate (MMS). Acetylation increases its binding affinity to the negative calcium response element (nCaRE) DNA promoter. The acetylated form induces a stronger binding of YBX1 to the Y-box sequence in the MDR1 promoter than the unacetylated form. Deacetylated on lysines. Lys-6 and Lys-7 are deacetylated by SIRT1.
Cleaved at Lys-31 by granzyme A to create the mitochondrial form; leading in reduction of binding to DNA, AP endodeoxynuclease activity, redox activation of transcription factors and to enhanced cell death. Cleaved by granzyme K; leading to intracellular ROS accumulation and enhanced cell death after oxidative stress.
Cys-65 and Cys-93 are nitrosylated in response to nitric oxide (NO) and lead to the exposure of the nuclear export signal (NES).
Ubiquitinated by MDM2; leading to translocation to the cytoplasm and proteasomal degradation.
细胞定位Mitochondrion. The cleaved APEX2 is only detected in mitochondria (By similarity). Translocation from the cytoplasm to the mitochondria is mediated by ROS signaling and cleavage mediated by granzyme A. Tom20-dependent translocated mitochondrial APEX1 level is significantly increased after genotoxic stress and Nucleus. Nucleus, nucleolus. Nucleus speckle. Endoplasmic reticulum. Cytoplasm. Detected in the cytoplasm of B-cells stimulated to switch (By similarity). Colocalized with SIRT1 in the nucleus. Colocalized with YBX1 in nuclear speckles after genotoxic stress. Together with OGG1 is recruited to nuclear speckles in UVA-irradiated cells. Colocalized with nucleolin and NPM1 in the nucleolus. Its nucleolar localization is cell cycle dependent and requires active rRNA transcription. Colocalized with calreticulin in the endoplasmic reticulum. Translocation from the nucleus to the cytoplasm is stimulated in presence of nitric oxide (NO) and function in a CRM1-dependent manner, possibly as a consequence of demasking a nuclear export signal (amino acid position 64-80). S-nitrosylation at Cys-93 and Cys-310 regulates its nuclear-cytosolic shuttling. Ubiquitinated form is localized predominantly in the cytoplasm.
- Information by UniProt
- AP endonuclease 1 antibody
- AP endonuclease class I antibody
- AP lyase antibody
All lanes : Anti-APE1 antibody [13B8E5C2] - ChIP Grade (ab194) at 1/2000 dilution
Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : APEX1 knockout HAP1 whole cell lysate
Lane 3 : HepG2 whole cell lysate
Lane 4 : HEK293 whole cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 35.6 kDa
Observed band size: 37 kDa why is the actual band size different from the predicted?
Lanes 1 - 4: Merged signal (red and green). Green - ab194 observed at 37 kDa. Red - loading control, ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) observed at 55kDa.
ab194 was shown to react with APEX1 in HAP1 wild-type cells in Western blot. Loss of signal was observed when APEX1 knockout sample was used. HAP1 wild-type and APEX1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% Milk in TBS-T (0.1% Tween®) before incubation with ab194 and ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) overnight at 4°C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
ab194 staining cultured human HeLa cells by ICC/IF. Cells were PFA fixed and permeabilized in 0.5% Triton X100 prior to blocking in 5% BSA for 1 hour at 20°C. The primary antibody was diluted 1/200 and incubated with the sample for 1 hour at 20°C. A Cy3® conjugated donkey anti-mouse antibody diluted was used as the secondary.
Ab194 staining Human normal placenta. Staining is localized to the nucleus.
Left panel: with primary antibody at 1 ug/ml. Right panel: isotype control.
Sections were stained using an automated system DAKO Autostainer Plus , at room temperature. Sections were rehydrated and antigen retrieved with the Dako 3-in-1 AR buffer citrate pH 6.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS), then incubated with primary antibody for 20 minutes, and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.
Anti-APE1 antibody [13B8E5C2] - ChIP Grade (ab194) at 1/2000 dilution + Human fibroblast at 30 µg with Milk at 5 %
ECL Mouse IgG,HRP - linked whole antibody (sheep) at 1/5000 dilution
Developed using the ECL technique.
Predicted band size: 35.6 kDa
Exposure time: 10 seconds
This product has been referenced in:
- de Jesus DS et al. Nox1/Ref-1-mediated activation of CREB promotes Gremlin1-driven endothelial cell proliferation and migration. Redox Biol 22:101138 (2019). Read more (PubMed: 30802716) »
- Kumar S et al. Role of apurinic/apyrimidinic nucleases in the regulation of homologous recombination in myeloma: mechanisms and translational significance. Blood Cancer J 8:92 (2018). Read more (PubMed: 30301882) »