Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-CD166 antibody [EPR2759(2)] - BSA and Azide free (ab206127)

Overview

  • Product name

    Anti-CD166 antibody [EPR2759(2)] - BSA and Azide free
    See all CD166 primary antibodies
  • Description

    Rabbit monoclonal [EPR2759(2)] to CD166 - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, IP, ICC/IF, Flow Cyt, IHC-Pmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human CD166. The exact sequence is proprietary.

  • Positive control

    • WB: SH-SY5Y, HuT-78, HT-1080 and Daudi cell lysates. Mouse and rat brain tissue lysates. IHC-P: Human liver and prostatic adenocarcinoma tissues. ICC/IF: Jurkat cells. Flow Cyt: HuT-78 cells. IP: SH-SY5Y cells.
  • General notes

    Ab206127 is the carrier-free version of ab109215. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    Use our conjugation kits  for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    ab206127 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab206127 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
WB Use at an assay dependent concentration. Predicted molecular weight: 65 kDa.

Please check the parent abID, ab109215, for more information on dilutions.

IP Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.

ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

 

IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

Target

  • Function

    Cell adhesion molecule that binds to CD6. Involved in neurite extension by neurons via heterophilic and homophilic interactions. May play a role in the binding of T- and B-cells to activated leukocytes, as well as in interactions between cells of the nervous system.
  • Tissue specificity

    Spleen, placenta, liver, and weakly in liver. Expressed by activated T-cells, B-cells, monocytes and thymic epithelial cells. Expressed by neurons in the brain. Restricted expression in tumor cell lines. Preferentially expressed in highly metastasizing melanoma cell lines.
  • Sequence similarities

    Contains 3 Ig-like C2-type (immunoglobulin-like) domains.
    Contains 2 Ig-like V-type (immunoglobulin-like) domains.
  • Domain

    The CD6 binding site is located in the N-terminal Ig-like domain.
  • Cellular localization

    Membrane.
  • Information by UniProt
  • Database links

  • Alternative names

    • Activated leukocyte cell adhesion molecule antibody
    • ALCAM antibody
    • ALCAM protein antibody
    • CD 166 antibody
    • CD166 antibody
    • CD166 antigen antibody
    • CD166_HUMAN antibody
    • FLJ3851 antibody
    • FLJ38514 antibody
    • MEMD antibody
    • MGC71733 antibody
    see all

Images

  • Flow Cytometry analysis of HuT-78 cells labelling CD166 with purified ab109215 at 1/90 (red). Cells were fixed with 2% paraformaldehyde. A FITC-conjugated goat anti-rabbit IgG (1/150) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109215).

  • Immunocytochemistry/Immunofluorescence analysis of THP-1 (human monocytic leukemia cell line) cells labelling CD166 (green) with purified ab109215 at 1/250. Cells were fixed with 100% methanol. ab150077, Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as a nuclear counterstain.  

    Secondary Only Control: PBS was used instead of the primary antibody as the negative control.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109215).

  • ab109215 (purified) at 1/30 immunoprecipitating CD166 in SH-SY5Y (human neuroblastoma cell line from bone marrow) cell lysate (Lane 1). Lane 2 - PBS. For western blotting, a HRP-conjugated anti-rabbit IgG, specific to the non-reduced form of IgG was used as the secondary antibody (1/1500).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109215).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human liver tissue labelling CD166 with purified ab109215 at 1/50. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. A prediluted HRP-polymer conjugated anti-rabbit IgG was used as the secondary antibody. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109215).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human liver tissue labelling CD166 with unpurified ab109215 at 1/100.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109215).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human prostatic adenocarcinoma tissue labelling CD166 with unpurified ab109215 at 1/100.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109215).

  • This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109215).

    Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
    Lane 2: CD166 knockout HAP1 whole cell lysate (20 µg)
    Lane 3: HeLa whole cell lysate (20 µg)
    Lane 4: SH-SY5Y whole cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab109215 observed at 100 kDa. Red - loading control, ab9484, observed at 37 kDa.

    ab109215 was shown to specifically react with CD166 in wild-type HAP1 cells as signal was lost in CD166 knockout cells. Wild-type and CD166 knockout samples were subjected to SDS-PAGE. Ab109215 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 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 1/20000 dilution for 1 hour at room temperature before imaging.

  • Anti-CD166 antibody [EPR2759(2)] - BSA and Azide free (ab206127) + SH-SY5Y (human neuroblastoma) whole cell lysate at 10 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051)

    Predicted band size: 65 kDa
    Observed band size: 100-105 kDa
    why is the actual band size different from the predicted?


    Exposure time: 1 minute


    Blocking buffer and concentration: 5% NFDM/TBST

    Diluting buffer and concentration: 5% NFDM/TBST

References

This product has been referenced in:

  • Zhao Y  et al. Activation of bone marrow-derived mesenchymal stromal cells-a new mechanism of defocused low-energy shock wave in regenerative medicine. Cytotherapy 15:1449-57 (2013). Flow Cyt ; Rat . Read more (PubMed: 24199590) »
See 1 Publication for this product

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