The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 5 µg/ml.
Use a concentration of 1 µg/ml. Detects a band of approximately 193 kDa (predicted molecular weight: 193 kDa).
E3 ubiquitin-protein ligase involved in DNA repair. Upon genotoxic stress, accepts ubiquitin from the UBE2N-UBE2V2 E2 complex and transfers it to 'Lys-164' of PCNA which had been monoubiquitinated by UBE2A/B-RAD18, promoting the formation of non-canonical poly-ubiquitin chains linked through 'Lys-63'.
ICC/IF image of ab80129 stained SHSY5Y cells. The cells were 4% formaldehyde 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 (ab80129, 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 result in 100% methanol fixed (5 min) SHSY5Y cells at 5µg/ml.
Western blot - Anti-SHPRH antibody (ab80129)
Anti-SHPRH antibody (ab80129) at 1 µg/ml + SHSY-5Y (Human neuroblastoma cell line) Whole Cell Lysate at 10 µg
Secondary Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
This antibody was raised against an immunogen that is predicted to recognize isoforms (1,2,3,4 and 5) of E3 ubiquitin-protein ligase SHPRH. The predicted molecular weights of isoforms (1,2,3,4 and 5) are 193 kDa, 104kDa, 17 kDa, 190 kDa and 122 kDa respectively.