Mark3 Antibody (JE44-84)

Western Blot: Mark3 Antibody (JE44-84) [NBP2-76884] - Western blot analysis of MARK3 on different lysates. Proteins were transferred to a PVDF membrane and blocked with 5% BSA in PBS for 1 hour at room temperature. The primary antibody was used at a 1:500 dilution in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:5,000 dilution was used for 1 hour at room temperature.Positive control: Lane 1: rat brain tissue lysateLane 2: A431 cell lysateLane 3: 293 cell lysate

Immunocytochemistry/Immunofluorescence: Mark3 Antibody (JE44-84) [NBP2-76884] - Immunocytochemistry staining MARK3 in SiHa cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with MARK3 monoclonal antibody at a dilution of 1:50 for 1 hour at room temperature, washed with PBS. Alexa Fluorc 488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/100 dilution. The nuclear counter stain is DAPI (blue).

Immunohistochemistry: Mark3 Antibody (JE44-84) [NBP2-76884] - Immunohistochemical analysis of paraffin-embedded mouse small intestine tissue using anti-MARK3 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes. The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the antibody at 1/100 dilution, for 30 minutes at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chrogen. Counter stained with hematoxylin and mounted with DPX.

Flow Cytometry: Mark3 Antibody (JE44-84) [NBP2-76884] - Flow cytometric analysis of MARK3 was done on MCF-7 cells. The cells were fixed, permeabilized and stained with MARK3 antibody at 1/100 dilution (red) compared with an unlabelled control (cells without incubation with primary antibody; black). After incubation of the primary antibody on room temperature for 1 hour, the cells was stained with a Alexa Fluor 488-conjugated goat anti-rabbit IgG Secondary antibody at 1/500 dilution for 30 minutes.

Immunocytochemistry/Immunofluorescence: Mark3 Antibody (JE44-84) [NBP2-76884] - Immunocytochemistry staining MARK3 in MCF-7 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with MARK3 monoclonal antibody at a dilution of 1:100 for 1 hour at room temperature, washed with PBS. Alexa Fluorc 488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/100 dilution. The nuclear counter stain is DAPI (blue).

Immunocytochemistry/Immunofluorescence: Mark3 Antibody (JE44-84) [NBP2-76884] - Immunocytochemistry staining MARK3 in SH-SY-5Y cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with MARK3 monoclonal antibody at a dilution of 1:50 for 1 hour at room temperature, washed with PBS. Alexa Fluorc 488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/100 dilution. The nuclear counter stain is DAPI (blue).

Immunohistochemistry: Mark3 Antibody (JE44-84) [NBP2-76884] - Immunohistochemical analysis of paraffin-embedded human appendix tissue using anti-MARK3 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes. The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the antibody at 1/100 dilution, for 30 minutes at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chrogen. Counter stained with hematoxylin and mounted with DPX.

Immunohistochemistry: Mark3 Antibody (JE44-84) [NBP2-76884] - Immunohistochemical analysis of paraffin-embedded human breast cancer tissue using anti-MARK3 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes. The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the antibody at 1/100 dilution, for 30 minutes at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chrogen. Counter stained with hematoxylin and mounted with DPX.

Product Discontinued

Mark3 Antibody (JE44-84) Summary

• Additional Information: Recombinant Monoclonal Antibody.
• Immunogen: Synthetic peptide within Human Mark3 aa 567-616 / 753. (SwissProt: P27448 Human; SwissProt: Q03141 Mouse; SwissProt: Q8VHF0 Rat)
• Localization: Cell membrane. Peripheral membrane protein.
• Isotype: IgG
• Clonality: Monoclonal
• Host: Rabbit
• Gene: MARK3
• Purity: Protein A purified

Applications/Dilutions

• Dilutions:
  • Flow Cytometry 1:50-1:100
  • Immunocytochemistry/ Immunofluorescence 1:50-1:200
  • Immunohistochemistry
  • Immunohistochemistry-Paraffin 1:50-1:200
  • Immunoprecipitation 1:10-1:50
  • Western Blot 1:500
• Theoretical MW: 84 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Packaging, Storage & Formulations

• Storage: Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.
• Buffer: TBS (pH7.4), 0.05% BSA, 40% Glycerol
• Preservative: 0.05% Sodium Azide
• Concentration: 1 mg/ml
• Purity: Protein A purified

Alternate Names for Mark3 Antibody (JE44-84)

• Cdc25C-associated protein kinase 1
• cTAK1
• C-TAK1
• CTAK1ELKL motif kinase 2
• EC 2.7.11
• EC 2.7.11.1
• EMK2
• EMK-2
• KP78
• MAP/microtubule affinity-regulating kinase 3
• PAR1A
• Protein kinase STK10
• Ser/Thr protein kinase PAR-1
• Serine/threonine-protein kinase p78

Background

MARK3 is a dual-specificity protein kinase that controls entry into mitosis by dephosphorylating Cdc2 on both threonine 14 and tyrosine 15. MARK3 is phosphorylated on serine 216 throughout interphase but not during mitosis. Serine 216 phosphorylation mediates the binding of 14-3-3 protein to MARK3, and MARK3/14-3-3 complexes are present throughout interphase but not during mitosis (1). Kinase suppressor of Ras (KSR) is a conserved component of the Ras pathway that interacts directly with MEK and MAPK. It has been shown that KSR1 translocates from the cytoplasm to the cell surface in response to growth factor treatment and that this process is regulated by MARK3 (2). MARK3 seems to be a positive regulator of the beta-catenin pathway and an inhibitor of the JNK pathway. These findings show that MARK3, a regulator of polarity, is also a modulator of Wnt-beta-catenin signalling, indicating a link between two important developmental pathways (3).

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Bioinformatics

• Gene Symbol: MARK3