This antibody gave a positive signal in the following lysates:
HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate
Heart (Mouse) Tissue Lysate
Testis (Mouse) Tissue Lysate
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The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml.
Use a concentration of 1 µg/ml. Detects a band of approximately 44 kDa (predicted molecular weight: 44 kDa).
Use at an assay dependent concentration. Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.
May play a role in a ERBB3-regulated signal transduction pathway. Seems be involved in growth regulation. Acts a corepressor of the androgen receptor (AR) and is regulated by the ERBB3 ligand neuregulin-1/heregulin (HRG). Inhibits transcription of some E2F1-regulated promoters, probably by recruiting histone acetylase (HAT) activity. Binds RNA. Associates with 28S, 18S and 5.8S mature rRNAs, several rRNA precursors and probably U3 small nucleolar RNA. May be involved in regulation of intermediate and late steps of rRNA processing. May be involved in ribosome assembly. Mediates cap-independent translation of specific viral IRESs (internal ribosomal entry site).
Expressed in several cell lines tested, including primary and transformed cell lines.
Belongs to the peptidase M24 family.
Phosphorylated on serine and threonine residues. Phosphorylation is enhanced by HRG treatment. Basal phosphorylation is PKC-dependent and HRG-induced phosphorylation is predominantly PKC-independent.
Cytoplasm. Nucleus > nucleolus. Tranlocates to the nucleus upon treatment with HRG.
This antibody detects a band corresponding to EBP1 at the predicted molecular weight of 44 kDa. An additional band of ~54 kDa is also detected. We are unsure of the identity of this band, although this may represent a modified form of the protein.
Our other antibody to this target (ab33613), raised against a distinct epitope, also detects the major band at 44 kDa. In contrast to ab35424 however, an additional band at 38 kDa is detected by ab33613. These distinct banding patterns suggest that this protein exists in multiple forms.
Western blot - EBP1 antibody (ab35424)
All lanes : Anti-EBP1 antibody (ab35424) at 1 µg/ml
Lane 1 : Heart (Mouse) Tissue Lysate Lane 2 : Testis (Mouse) Tissue Lysate
Lysates/proteins at 10 µg per lane.
Secondary All lanes : IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution
Performed under reducing conditions.
Predicted band size: 44 kDa Observed band size: 44 kDa Additional bands at: 55 kDa, 70 kDa. We are unsure as to the identity of these extra bands.
ICC/IF image of ab35424 stained human HeLa cells. The cells were PFA fixed (10 min), permabilised in TBS-T (20 min) and incubated with the antibody (ab35424, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal serum / 0.3M glycine was used to quench autofluorescence and block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue).
ab35424 staining in human kidney, showing staining of the tubuli cells (in brown). Paraffin embedded kidney tissue was incubated with ab35424 (1:600 dilution) for 30 mins at room temperature. Antigen retrieval was performed by heat induction in citrate buffer pH 6.
ab35424 was tested in a tissue microarray (TMA) containing a wide range of normal and cancer tissues as well as a cell microarray consisting of a range of commonly used, well characterised human cell lines. Further results for this antibody can be found at www.proteinatlas.org