The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml. Detects a band of approximately 125 kDa (predicted molecular weight: 123 kDa).
Use a concentration of 5 µg/ml.
Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Protein kinase that regulates key processes linked to cell growth and survival. Regulates cytoskeleton remodeling during cell differentiation, cell division and cell adhesion. Localizes to dynamic actin structures, and phosphorylates CRK and CRKL, DOK1, and other proteins controlling cytoskeleton dynamics. Regulates DNA repair potentially by activating the proapoptotic pathway when the DNA damage is too severe to be repaired. Phosphorylates PSMA7 that leads to an inhibition of proteasomal activity and cell cycle transition blocks.
Note=A chromosomal aberration involving ABL1 is a cause of chronic myeloid leukemia. Translocation t(9;22)(q34;q11) with BCR. The translocation produces a BCR-ABL found also in acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL).
Belongs to the protein kinase superfamily. Tyr protein kinase family. ABL subfamily. Contains 1 protein kinase domain. Contains 1 SH2 domain. Contains 1 SH3 domain.
Phosphorylated by PRKDC (By similarity). DNA damage-induced activation of c-Abl requires the function of ATM and Ser-446 phosphorylation (By similarity). Phosphorylation on Thr-735 is required for binding 14-3-3 proteins for cytoplasmic translocation. Isoform IB is myristoylated on Gly-2.
Cytoplasm > cytoskeleton. Nucleus. Sequestered into the cytoplasm through interaction with 14-3-3 proteins and Nucleus membrane. The myristoylated c-ABL protein is reported to be nuclear.
IHC image of Abl staining in mouse uterus formalin fixed paraffin embedded tissue section, performed on a Leica Bond system using the standard protocol B. 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 ab85947, 1µg/ml, for 15 mins at room temperature. A goat anti-rabbit biotinylated secondary antibody was used to detect the primary, and visualized using an HRP conjugated ABC 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.
Western blot - c Abl antibody (ab85947)
All lanes : Anti-c Abl antibody (ab85947) at 1 µg/ml
Lane 1 : Human breast tissue lysate - total protein (ab30090) Lane 2 : MCF7 (Human breast adenocarcinoma cell line) Whole Cell Lysate Lane 3 : Human ovary tissue lysate - total protein (ab30222) Lane 4 : K562 (Human erythromyeloblastoid leukemia cell line) Whole Cell Lysate Lane 5 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
Lysates/proteins at 10 µg per lane.
Secondary All lanes : Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
Immunocytochemistry/ Immunofluorescence - c Abl antibody (ab85947)
ICC/IF image of ab85947 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 (ab85947, 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.