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 28 kDa (predicted molecular weight: 30 kDa).
Enkephalin and enkephalin-like peptides have been detected in many animal species. In mammals these opioid pentapeptides are cleaved from a common precursor molecule which is expressed in a number of cell types, including neurons and endocrine cells associated with the sympathetic nervous system where they are often co-stored with catecholamines. They are known to be endogenous ligands for Sopioid receptors and are known to mediate analgesia at the spinal level.
The band observed at 28 kDa could potentially be a cleaved form of Leu Enkephalin due to the presence of a 24 amino acid signal peptide.
Immunocytochemistry/ Immunofluorescence - Leu Enkephalin antibody (ab85798)
ICC/IF image of ab85978 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 (ab85798, 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 4% PFA fixed (10 min) Hek293, HepG2 and MCF7 cells at 5µg/ml, and in 100% methanol fixed (5 min) HeLa, Hek293, HepG2 and MCF7 cells at 5µg/ml.