Overview

  • Product name

    Anti-CLC3 antibody [N258/5]
    See all CLC3 primary antibodies
  • Description

    Mouse monoclonal [N258/5] to CLC3
  • Host species

    Mouse
  • Specificity

    Does not cross-react with CLCN4 or CLCN5 (based on KO validation results).
  • Tested applications

    Suitable for: Flow Cyt, WB, IHC-P, ICC/IFmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide corresponding to Rat CLC3 aa 98-115 (N terminal).
    Sequence:

    CKDRERHRRINSKKKESA


    Database link: NP_445815.2

  • Positive control

    • WB: Rat brain membrane lysate. IHC-P: FFPE human hippocampus tissue sections. ICC/IF: SK-N-BE cells.
  • General notes

    The clone number has been updated from S258-5 to N258/5, both clone numbers name the same antibody clone.

     

    Previously labelled as CLCN3. 

Properties

Applications

Our Abpromise guarantee covers the use of ab134285 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
Flow Cyt Use 1µg for 106 cells.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

 

WB 1/1000. Predicted molecular weight: 91 kDa.

1µg/ml of ab134285 is sufficient for detection of CLC3 in 20µg of Rat brain membrane lysate.

IHC-P 1/1000.
ICC/IF 1/100.

Target

  • Function

    Mediates the exchange of chloride ions against protons. Functions as antiporter and contributes to the acidification of the endosome and synaptic vesicle lumen, and may thereby affect vesicle trafficking and exocytosis. May play an important role in neuronal cell function through regulation of membrane excitability by protein kinase C. It could help neuronal cells to establish short-term memory.
  • Tissue specificity

    Expressed primarily in tissues derived from neuroectoderm. Within the brain, its expression is particularly evident in the hippocampus, olfactory cortex, and olfactory bulb. Highly expressed in aortic and coronary vascular smooth muscle cells, and aortic endothelial cells. Also expressed in tracheal and alveolar epithelial cells, and intima and media of the pulmonary vessels. Expressed in bronchus and colon (at protein level).
  • Sequence similarities

    Belongs to the chloride channel (TC 2.A.49) family. ClC-3/CLCN3 subfamily.
    Contains 2 CBS domains.
  • Domain

    Isoform 2 contains a C-terminal PDZ-binding motif mediating the interaction with GOPC.
  • Post-translational
    modifications

    N-glycosylated.
  • Cellular localization

    Membrane. Early endosome membrane. Late endosome membrane. Cytoplasmic vesicle > secretory vesicle membrane. Isoform 1 is localized mainly in early and late endosomes and Membrane. Early endosome membrane. Late endosome membrane. Golgi apparatus membrane. Isoform 2 partially colocalized with isoform 1 but is mainly enriched in the Golgi.
  • Information by UniProt
  • Database links

  • Alternative names

    • Chloride channel 3 antibody
    • Chloride channel protein 3 antibody
    • Chloride channel, voltage-sensitive 3 antibody
    • Chloride transporter ClC 3 antibody
    • Chloride transporter ClC-3 antibody
    • Chloride transporter ClC3 antibody
    • chloride voltage-gated channel 3 antibody
    • ClC 3 antibody
    • ClC-3 antibody
    • ClC3 antibody
    • CLCN 3 antibody
    • CLCN3 antibody
    • CLCN3_HUMAN antibody
    • DKFZp564I0463 antibody
    • H(+)/Cl(-) exchange transporter 3 antibody
    see all

Images

  • Anti-CLC3 antibody [N258/5] (ab134285) at 1/200 dilution + Rat brain membrane lysate at 15 µg

    Secondary
    Goat Anti-Mouse IgG: HRP at 1/1000 dilution

    Developed using the ECL technique.

    Predicted band size: 91 kDa

  • 4% formaldehyde-fixed SK-N-BE cells stained for CLC3 (green) using ab134285 at 1/100 dilution in ICC/IF.

    Secondary Antibody: Goat Anti-Mouse ATTO 488 at 1/100 for 60 minutes at room temperature. Counterstain: Phalloidin Texas Red F-Actin stain; DAPI (blue) nuclear stain at 1/1000, 1/5000 for 60 minutes room temperature, 5 minutes room temperature.

    (A) DAPI (blue) nuclear stain. (B) Phalloidin Texas Red F-Actin stain. (C) ab134285. (D) Composite.

  • IHC image of CLC3 staining in human hippocampus formalin fixed paraffin embedded tissue section, performed on a Leica Bond system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab134285, 10µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • Overlay histogram showing SH-SY5Y cells stained with ab134285 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab134285, 1μg/1x106 cells) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150113) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

References

This product has been referenced in:

See 1 Publication for this product

Customer reviews and Q&As

Answer

Thank you for contacting us.

We currently have three anti-CLCN3 products on our catalog. Unfortunately these are not phospho specific antibody and will recognize CLCN3 regardless of phosphorylation state. I can suggest two courses of action for detection of phosphorylated CLCN3. The first is to have a custom antibody created to specifically detected CLCN3 phosphorylated at S109. Our Epitomics group does provide custom rabbit monoclonal antibody services. Rabbits have a unique immune system which allows them to develop antibodies against “small epitopes” present in small molecules, lipids and polymers or subtle changes in epitopes such as post-translational modifications or single amino acid substitution. Rabbit monoclonal offer superior affinity as well. More information about our custom antibodies services may be found at: http://www.epitomics.com/services/ or by emailing service@epitomics.com.

You may also choose to use a non phospho specific antibody such as https://www.abcam.com/CIDE-C-antibody-ab77115.html, https://www.abcam.com/CIDE-C-antibody-ab16760.html or https://www.abcam.com/CIDE-C-antibody-ab102016.html in a WB system which incorporates a phos-tag acrylamide. These reagents specifically bind phosphate groups and can be incorporated into SDS-PAGE gels. These bind to phosphorylated proteins and hamper their progress through the gel. The more a protein is phosphorylated, the slower it will migrate and should resolve different phospho-species as individual bands. While this is an interesting system, we have not yet tested the phos-tag system and cannot guarantee its effectiveness.


I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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