Recombinant Anti-CD9 antibody [EPR23105-121] (ab236630)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR23105-121] to CD9
- Suitable for: WB, ICC/IF, IHC-P, Flow Cyt, IP
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
Overview
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Product name
Anti-CD9 antibody [EPR23105-121]
See all CD9 primary antibodies -
Description
Rabbit monoclonal [EPR23105-121] to CD9 -
Host species
Rabbit -
Tested applications
Suitable for: WB, ICC/IF, IHC-P, Flow Cyt, IPmore details -
Species reactivity
Reacts with: Human -
Immunogen
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: HeLa, MCF7, A549, HCT 116, Human tonsil and Human colon lysates. IHC-P: Human spleen and Human bladder carcinoma tissues. ICC/IF: HCT 116 cells. Flow Cyt: HCT 116 cells. IP: HCT 116 cell lysate.
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General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR23105-121 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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KO cell lines
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KO cell lysates
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Recombinant Protein
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab236630 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 |
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WB | (2) |
1/1000. Detects a band of approximately 22 kDa (predicted molecular weight: 25 kDa).
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ICC/IF | (1) |
1/500.
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IHC-P |
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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Flow Cyt |
1/50.
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IP |
1/30.
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Notes |
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WB
1/1000. Detects a band of approximately 22 kDa (predicted molecular weight: 25 kDa).
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ICC/IF
1/500. |
IHC-P
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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Flow Cyt
1/50.
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IP
1/30.
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Target
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Function
Involved in platelet activation and aggregation. Regulates paranodal junction formation. Involved in cell adhesion, cell motility and tumor metastasis. Required for sperm-egg fusion. -
Tissue specificity
Expressed by a variety of hematopoietic and epithelial cells. -
Sequence similarities
Belongs to the tetraspanin (TM4SF) family. -
Post-translational
modificationsProtein exists in three forms with molecular masses between 22 and 27 kDa, and is known to carry covalently linked fatty acids. -
Cellular localization
Membrane. - Information by UniProt
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Database links
- Entrez Gene: 928 Human
- Omim: 143030 Human
- SwissProt: P21926 Human
- Unigene: 114286 Human
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Alternative names
- Tetraspanin 29 antibody
- 5H9 antibody
- 5H9 antigen antibody
see all
Images
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All lanes : Anti-CD9 antibody [EPR23105-121] (ab236630) at 1/1000 dilution
Lane 1 : Wild-type HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : CD9 knockout HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 3 : A549 (Human lung carcinoma cell line) whole cell lysate
Lane 4 : MCF7 (Human breast adenocarcinoma cell line) whole cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 25 kDa
Observed band size: 18 kDa why is the actual band size different from the predicted?Lanes 1 - 4: Merged signal (red and green). Green - ab236630 observed at 18 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.
ab236630 was shown to react with CD9 in wild-type HeLa cells in Western blot with loss of signal observed in CD9 knockout cell line ab255375 (CD9 knockout cell lysate ab263754). Wild-type HeLa and CD9 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab236630 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
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Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HCT 116 (human colorectal carcinoma epithelial cell) cells labelling CD9 with ab236630 at 1/500 dilution, followed by ab150077 AlexaFluor®488 Goat anti-Rabbit secondary antibody at 1/1000 dilution (Green). Confocal image showing membranous staining in HCT 116 cell line. Negative control: Raji (PMID: 8921952). ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is ab150077 AlexaFluor®488 Goat anti-Rabbit secondary at 1/1000 dilution.
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All lanes : Anti-CD9 antibody [EPR23105-121] (ab236630) at 1/1000 dilution
Lane 1 : Human tonsil lysate
Lane 2 : Human colon lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 25 kDa
Observed band size: 22 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 5% NFDM/TBST. The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 3275469). Exposure time: 3 minutes.
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All lanes : Anti-CD9 antibody [EPR23105-121] (ab236630) at 1/1000 dilution
Lane 1 : HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate
Lane 2 : Raji (human Burkitts lymphoma b lymphocyte) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 25 kDa
Observed band size: 22 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 5% NFDM/TBST This blot was developed using a higher sensitivity ECL substrate. The molecular weight observed is consistent with what has been described in the literature (PMID: 3275469).
Negative control: Raji (PMID: 8921952). Exposure time: 3 minutes
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CD9 was immunoprecipitated from 0.35 mg HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate with ab236630 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab236630 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.
Lane 1: HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate 10ug.
Lane 2: ab236630 IP in HCT 116 whole cell lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab236630 in HCT 116 whole cell lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 30 seconds.
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Flow cytometric analysis of Raji (human Burkitt's lymphoma B lymphocyte, Left) / HCT 116 (human colorectal carcinoma epithelial cell, Right) cells labelling CD9 with ab236630 at 1/50 dilution compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 was used as the secondary antibody.
Negative control: Raji (PMID: 8921952).
Gated on viable cells.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD9 antibody [EPR23105-121] (ab236630)
Immunohistochemical analysis of paraffin-embedded Human bladder carcinoma tissue labeling CD9 with ab236630 at 1/100 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on human bladder carcinoma. The section was incubated with ab236630 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD9 antibody [EPR23105-121] (ab236630)
Immunohistochemical analysis of paraffin-embedded Human spleen tissue labeling CD9 with ab236630 at 1/100 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on platelets of human spleen. The section was incubated with ab236630 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins.
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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SDS download
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Datasheet download
Certificate of Compliance
References (2)
ab236630 has been referenced in 2 publications.
- Wang S et al. Transfer of microRNA-25 by colorectal cancer cell-derived extracellular vesicles facilitates colorectal cancer development and metastasis. Mol Ther Nucleic Acids 23:552-564 (2021). PubMed: 33510943
- Mathieu M et al. Specificities of exosome versus small ectosome secretion revealed by live intracellular tracking of CD63 and CD9. Nat Commun 12:4389 (2021). PubMed: 34282141