The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
1/250. Detects a band of approximately 120 kDa (predicted molecular weight: 110 kDa).
Use a concentration of 5 µg/ml.
FunctionDocking protein which plays a central coordinating role for tyrosine-kinase-based signaling related to cell adhesion. May function in transmitting growth control signals between focal adhesions at the cell periphery and the mitotic spindle in response to adhesion or growth factor signals initiating cell proliferation. May play an important role in integrin beta-1 or B cell antigen receptor (BCR) mediated signaling in B- and T-cells. Integrin beta-1 stimulation leads to recruitment of various proteins including CRK, NCK and SHPTP2 to the tyrosine phosphorylated form.
Tissue specificityWidely expressed. Higher levels detected in kidney, lung, and placenta. Also detected in T-cells, B-cells and diverse cell lines. The protein has been detected in lymphocytes, in diverse cell lines, and in lung tissues.
Sequence similaritiesBelongs to the CAS family. Contains 1 SH3 domain.
DomainContains a central domain containing multiple potential SH2-binding sites and a C-terminal domain containing a divergent helix-loop-helix (HLH) motif. The SH2-binding sites putatively bind CRK, NCK and ABL SH2 domains. The HLH motif confers specific interaction with the HLH proteins ID2, E12 and E47. It is absolutely required for the induction of pseudohyphal growth in yeast and mediates homodimerization and heterodimerization with p130cas. The SH3 domain interacts with two proline-rich regions of focal adhesion kinase.
Post-translational modificationsCell cycle-regulated processing produces four isoforms: p115, p105, p65, and p55. Isoform p115 arises from p105 phosphorylation and appears later in the cell cycle. Isoform p55 arises from p105 as a result of cleavage at a caspase cleavage-related site and it appears specifically at mitosis. The p65 isoform is poorly detected. Focal adhesion kinase 1 phosphorylates the protein at the YDYVHL motif (conserved among all cas proteins). The SRC family kinases (FYN, SRC, LCK and CRK) are recruited to the phosphorylated sites and can phosphorylate other tyrosine residues. Ligation of either integrin beta-1 or B-cell antigen receptor on tonsillar B-cells and B-cell lines promotes tyrosine phosphorylation and both integrin and BCR-mediated tyrosine phosphorylation requires an intact actin network. In fibroblasts transformation with oncogene v-ABL results in an increase in tyrosine phosphorylation. Transiently phosphorylated following CD3 cross-linking and this phosphorylated form binds to CRK and C3G. A mutant lacking the SH3 domain is phosphorylated upon CD3 cross-linking but not upon integrin beta-1 cross-linking. Tyrosine phosphorylation occurs upon stimulation of the G-protein coupled C1a calcitonin receptor in rabbit. Calcitonin-stimulated tyrosine phosphorylation is mediated by calcium- and protein kinase C-dependent mechanisms and requires the integrity of the actin cytoskeleton.
Cellular localizationCytoplasm > cytoskeleton > spindle and Cytoplasm > cell cortex. Nucleus. Golgi apparatus. Cell projection > lamellipodium. Cytoplasm. Cell junction > focal adhesion. Localizes to both the cell nucleus and the cell periphery and is differently localized in fibroblasts and epithelial cells. In fibroblasts is predominantly nuclear and in some cells is present in the Golgi apparatus. In epithelial cells localized predominantly in the cell periphery with particular concentration in lamellipodia but is also found in the nucleus. Isoforms p105 and p115 are predominantly cytoplasmic and associate with focal adhesions while p55 associates with mitotic spindle.
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Anti-HEF1 antibody images
Western blot - HEF1 antibody (ab37161)
All lanes : Anti-HEF1 antibody (ab37161) at 1/250 dilution
Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate Lane 2 : Jurkat whole cell lysate (ab7899) Lane 3 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate with Human HEF1 peptide (ab39117) at 1 µg/ml Lane 4 : Jurkat whole cell lysate (ab7899) with Human HEF1 peptide (ab39117) at 1 µg/ml
Immunocytochemistry/ Immunofluorescence - HEF1 antibody (ab37161)This image is courtesy of an Anonymous Abreview.
ab37161 staining HEF1 in Human glioblastoma cells by Immunocytochemistry/ Immunofluorescence. Cells were PFA-fixed and permeabilized in 0.1% Triton X-100 in PBS prior to blocking in 0.5% BSA in TBS-Tween for 20 minutes at room temperature. The primary antibody was diluted 1/50 in 0.5% BSA/PBS and incubated with the sample for 16 hours at 4°C. The secondary antibody was Cy3®-conjugated Goat anti-Rabbit IgG, diluted 1/400. Nuclei were counterstained with Hoechst.
ICC/IF image of ab37161 stained HeLa cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab37161, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899 Dylight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
References for Anti-HEF1 antibody (ab37161)
This product has been referenced in:
Knutson DC et al. Mice null for NEDD9 (HEF1a) display extensive hippocampal dendritic spine loss and cognitive impairment. Brain Res1632:141-55 (2016).
Read more (PubMed: 26683084) »