Overview

  • Product name

    Mouse APE ELISA Kit
    See all APE1 kits
  • Detection method

    Colorimetric
  • Precision

    Intra-assay
    Sample n Mean SD CV%
    C2C12 8 3.1%
    Inter-assay
    Sample n Mean SD CV%
    C2C12 3 8.8%
  • Sample type

    Cell culture supernatant, Cell culture extracts, Tissue Extracts
  • Assay type

    Sandwich (quantitative)
  • Sensitivity

    28.8 pg/ml
  • Range

    46.88 pg/ml - 3000 pg/ml
  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Cell culture media 100 98% - 104%

  • Assay time

    1h 30m
  • Assay duration

    One step assay
  • Species reactivity

    Reacts with: Mouse
  • Product overview

    Abcam’s mouse APE (Apex1) in vitro SimpleStep ELISA® (Enzyme-Linked Immunosorbent Assay) kit is designed for the quantitative measurement of mouse APE protein in mouse cell culture supernatants and cell and tissue extracts.

    The SimpleStep ELISA® employs an affinity tag labeled capture antibody and a reporter conjugated detector antibody which immunocapture the sample analyte in solution. This entire complex (capture antibody/analyte/detector antibody) is in turn immobilized via immunoaffinity of an anti-tag antibody coating the well. To perform the assay, samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material. TMB substrate is added and during incubation is catalyzed by HRP, generating blue coloration. This reaction is then stopped by addition of Stop Solution completing any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm. 

    Sensitivity:
    Samples diluted in Sample Diluent NS – 28.8 pg/mL
    Samples diluted in 1X Cell Extraction Buffer PTR – 4.6 pg/mL

  • Notes

    Mouse APE is a multifunctional protein that binds DNA and RNA and plays a central role in the cellular response to oxidative stress. The two major activities of APE are DNA repair and redox regulation of transcription factors. APE functions as an apurinic/apyrimidinic (AP) endodeoxyribonuclease in the DNA base excision repair (BER) pathway of DNA lesions induced by oxidative and alkylating agents. The C-terminal region of APE is responsible for base excision repair activities. Notably, this repair activity is suppressed by phosphorylation. The N-terminal region of APE may regulate the DNA binding capabilities of a variety of transcription factors through redox stimulation. Mouse APE has 98% and 94% identity to rat and human APE, respectively.

  • Tested applications

    Suitable for: Sandwich ELISAmore details
  • Platform

    Microplate (12 x 8 well strips)

Properties

  • Storage instructions

    Store at +4°C. Please refer to protocols.
  • Components 1 x 96 tests
    10X Mouse APE Capture Antibody 1 x 600µl
    10X Mouse APE Detector Antibody 1 x 600µl
    10X Wash Buffer PT (ab206977) 1 x 20ml
    50X Cell Extraction Enhancer Solution (ab193971) 1 x 1ml
    5X Cell Extraction Buffer PTR (ab193970) 1 x 10ml
    ab221824 - Antibody Diluent 4BI 1 x 6ml
    Mouse APE Lyophilized Recombinant Protein 2 vials
    Plate Seals 1 unit
    Sample Diluent NS 1 x 50ml
    SimpleStep Pre-Coated 96-Well Microplate (ab206978) 1 unit
    Stop Solution 1 x 12ml
    TMB Development Solution 1 x 12ml
  • Research areas

  • Function

    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.
  • Sequence similarities

    Belongs to the DNA repair enzymes AP/ExoA family.
  • Domain

    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.
  • Post-translational
    modifications

    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.
  • Cellular localization

    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
  • Alternative names

    • AP endonuclease 1
    • AP endonuclease class I
    • AP lyase
    • APE
    • APE 1
    • APE-1
    • APEN
    • APEX
    • APEX 1
    • APEX nuclease
    • APEX nuclease (multifunctional DNA repair enzyme) 1
    • Apex nuclease 1
    • APEX1
    • APEX1_HUMAN
    • Apurinic endonuclease
    • Apurinic-apyrimidinic endonuclease 1
    • Apurinic/apyrimidinic (abasic) endonuclease
    • Apurinic/apyrimidinic endonuclease 1
    • Apurinic/apyrimidinic exonuclease
    • APX
    • BAP1
    • Deoxyribonuclease (apurinic or apyrimidinic)
    • DNA (apurinic or apyrimidinic site) lyase
    • DNA-(apurinic or apyrimidinic site) lyase, mitochondrial
    • EC 4.2.99.18
    • HAP 1
    • HAP1
    • Human Apurinic endonuclease 1
    • MGC139790
    • Multifunctional DNA repair enzyme
    • Redox factor 1
    • Redox factor-1
    • REF 1
    • REF 1 protein
    • REF-1
    • REF1
    • REF1 protein
    see all
  • Database links

Associated products

Applications

Our Abpromise guarantee covers the use of ab207616 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
Sandwich ELISA Use at an assay dependent concentration.

Images

  • SimpleStep ELISA technology allows the formation of the antibody-antigen complex in one single step, reducing assay time to 90 minutes. Add samples or standards and antibody mix to wells all at once, incubate, wash, and add your final substrate. See protocol for a detailed step-by-step guide.

     

  • Background-subtracted data values (mean +/- SD) are graphed.

  • Background-subtracted data values (mean +/- SD) are graphed.

  • The concentrations of APE were measured in duplicate and interpolated from the APE standard curve and corrected for sample dilution. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean APE concentration was determined to be 3.26 ng/mL in C2C12 cell extract and 0.55 ng/mL in NIH 3T3 cell extract.

  • The concentrations of APE were measured in duplicate and interpolated from the APE standard curve and corrected for sample dilution. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean APE concentration was determined to be 2.09 ng/mL in rat PC12 cell extract and 2.84 ng/mL in mouse PC3 cell extract.

  • The concentrations of APE were measured in duplicates, interpolated from the APE standard curve and corrected for sample dilution. Undiluted samples are as follows: RPMI Base Media 10%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean APE concentration was determined to be 1,616 pg/mL in RPMI Base Media 10%.

Protocols

References

ab207616 has not yet been referenced specifically in any publications.

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