APE1 Mouse Monoclonal Antibody [12H2]
Catalog# EM1801-12
APE1 Mouse Monoclonal Antibody [12H2]
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WB
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IF-Cell
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IHC-P
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FC
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Human
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Mouse
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Rat
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unconjugated
Overview
Product Name
APE1 Mouse Monoclonal Antibody [12H2]
Antibody Type
Mouse Monoclonal Antibody
Immunogen
Recombinant protein within Human APE1 aa 20-318/318.
Species Reactivity
Human, Mouse, Rat
Validated Applications
WB, IF-Cell, IHC-P, FC
Molecular Weight
Predicted band size: 36 kDa
Positive Control
HeLa cell lysate, LNCaP cell lysate, HEK-293 cell lysate, HL-60 cell lysate, A431 cell lysate, NIH/3T3 cell lysate, mouse brain tissue lysate, C6 cell lysate, rat testis tissue lysate, A549, human kidney tissue, mouse liver tissue, rat colon tissue, HeLa.
Conjugation
unconjugated
Clone Number
12H2
RRID
Product Features
Form
Liquid
Concentration
Storage Instructions
Shipped at 4℃. Store at +4℃ short term (1-2 weeks). It is recommended to aliquot into single-use upon delivery. Store at -20℃ long term.
Storage Buffer
1*PBS (pH7.4), 0.2% BSA, 50% Glycerol. Preservative: 0.05% Sodium Azide.
Isotype
IgG2b
Purification Method
Immunogen affinity purified.
Application Dilution
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WB
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1:1,000-1:5,000
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IF-Cell
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1:100
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IHC-P
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1:1,000
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FC
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1:1,000
Target
Function
The role of transcription factors in the regulation of gene expression is well established. Although the activity of these factors can be regulated by phosphorylation, evidence has indicated regulation of DNA binding mediated by changes in reduction-oxidation (redox) status. Mutational analysis has identified a single conserved cysteine residue mapping within the DNA binding domains of Fos and Jun. Chemical oxidation or modification of this cysteine residue inhibits the DNA binding activity of Fos and Jun. A similar mode of regulation has been recently proposed for other nuclear transcription factors. Oxidation is reversible by these compounds or by a cellular redox/DNA repair protein identified originally as Ref-1 (redox factor 1). Ref-1 is identical to a previously characterized DNA repair enzyme designated HAP1, APE or APEX.
Background References
1. Robson C N et al. Isolation of cDNA clones encoding a human apurinic/apyrimidinic endonuclease that corrects DNA repair and mutagenesis defects in E. coli xth (exonuclease III) mutants. Nucleic Acids Res 19:5519-5523 (1991).
2. Fan Z et al. Cleaving the oxidative repair protein Ape1 enhances cell death mediated by granzyme A. Nat Immunol 4:145-153 (2003).
Sequence Similarity
Belongs to the DNA repair enzymes AP/ExoA family.
Post-translational Modification
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.
Subcellular Location
Nucleus. Endoplasmic reticulum, Cytoplasm.
Synonyms
AP endonuclease 1 antibody
AP endonuclease class I antibody
AP lyase antibody
APE 1 antibody
APE antibody
APE-1 antibody
APEN antibody
APEX 1 antibody
APEX antibody
APEX nuclease (multifunctional DNA repair enzyme) 1 antibody
ExpandImages
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Western blot analysis of APE1 on different lysates with Mouse anti-APE1 antibody (EM1801-12) at 1/1,000 dilution.
Lane 1: HeLa cell lysate (20 µg/Lane)
Lane 2: LNCaP cell lysate (20 µg/Lane)
Lane 3: HEK-293 cell lysate (20 µg/Lane)
Lane 4: HL-60 cell lysate (20 µg/Lane)
Lane 5: A431 cell lysate (20 µg/Lane)
Lane 6: NIH/3T3 cell lysate (20 µg/Lane)
Lane 7: Mouse brain tissue lysate (40 µg/Lane)
Lane 8: C6 cell lysate (20 µg/Lane)
Lane 9: Rat testis tissue lysate (40 µg/Lane)
Predicted band size: 36 kDa
Observed band size: 36 kDa
Exposure time: 1 minute;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (EM1801-12) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Mouse IgG - HRP Secondary Antibody (HA1006) at 1/50,000 dilution was used for 1 hour at room temperature. -
☑ Knockdown (KD)
Western blot analysis of APE1 on different lysates with Mouse anti-APE1 antibody (EM1801-12) at 1/5,000 dilution.
Lane 1:TE1-parental cell lysate
Lane 2: TE1-APE1 KD cell lysate
Lysates/proteins at 10 µg/Lane.
Predicted band size: 36 kDa
Observed band size: 36 kDa
Exposure time: 15 seconds; ECL: K1801;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (EM1801-12)) at 1/5,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature. -
Immunocytochemistry analysis of A549 cells labeling APE1 with Mouse anti-APE1 antibody (EM1801-12) at 1/100 dilution.
Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Mouse anti-APE1 antibody (EM1801-12) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Mouse IgG H&L (iFluor™ 488, HA1125) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
beta Tubulin (ET1602-4, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Rabbit IgG H&L (iFluor™ 594, HA1122) were used as the secondary antibody at 1/1,000 dilution. -
Immunohistochemical analysis of paraffin-embedded human kidney tissue with Mouse anti-APE1 antibody (EM1801-12) at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (EM1801-12) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
Immunohistochemical analysis of paraffin-embedded mouse liver tissue with Mouse anti-APE1 antibody (EM1801-12) at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (EM1801-12) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
Immunohistochemical analysis of paraffin-embedded rat colon tissue with Mouse anti-APE1 antibody (EM1801-12) at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (EM1801-12) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
Flow cytometric analysis of HeLa cells labeling APE1.
Cells were fixed and permeabilized. Then stained with the primary antibody (EM1801-12, 1μg/mL) (red) compared with Mouse IgG1 Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Mouse IgG Secondary antibody (HA1125) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"