This antibody gave a positive result in ELISA against the immunizing peptide (ab74478).
IF/ICC: See image below
IHC-P: See image below
Not yet tested in other applications.
GAPex 5 plays a role in several processes such as endocytosis, insulin receptor internalisation and LC2A4/GLUT4 trafficking. It acts as both a GTPase-activating protein (GAP) and a guanine nucleotide exchange factor (GEF). GAPex 5 is a GEF for the small G protein Rab31, promoting the exchange of GDP bound to Rab31 for GTP and thus regulating LC2A4/GLUT4 trafficking. In the absence of insulin, GAPex 5 maintains Rab31 in an active (GTP-bound) state, thereby promoting a futile cycle between LC2A4/GLUT4 storage vesicles and early endosomes, keeping LC2A4/GLUT4 inside the cells. Following insulin stimulation, GAPex 5 is translocated to the plasma membrane, enabling the release of LC2A4/GLUT4 from the intracellular storage vesicles. GAPex 5 is also involved in EGFR trafficking and degradation, and also has GEF activity for Rab5 and GAP activity for Ras.
Cell Membrane (peripheral membrane protein) and Cytoplasmic
ICC/IF image of ab65929 stained MCF7 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 (ab65929, 5µg/ml) overnight at +4°C. 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) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.This antibody also gave a positive IF result in xxxx cells. However, this Fast-Track antibody is not yet fully characterised. This image represents inconclusive preliminary data.
IHC image of ab65929 staining in human breast carcinoma formalin fixed paraffin embedded tissue section, performed on a Leica BondTM 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 ab65929, 5µg/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.
has not yet been referenced specifically in any publications.