Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis human hepatocarcinoma tissue sections labeling Alkaline Phosphatase with ab75699 at 1/25. Tissue was fixed with formaldehyde and blocked with 3% BSA for 0.5 hour at 37°C. Heat mediated antigen retrieval was performed using a citrate buffer (pH6). Samples were incubated with primary antibody (1/25) for 1 hour at 37°C. A Peroxidase-conjugated Goat anti-rabbit polyclonal (ready to use) was used as the secondary antibody.
Western blot - Anti-Alkaline Phosphatase antibody (ab75699)
Anti-Alkaline Phosphatase antibody (ab75699) at 1 µg/ml + Human liver tissue lysate - total protein (ab29889) at 10 µg
Secondary Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution Developed using the ECL technique
Exposure time : 2 minutesAlkaline Phosphatase protein contains a 22-amino acid signal peptide and a 29-amino acid propeptide, both of which are removed in the mature form (SwissProt). This might explain the migration of the band at a lower molecular weight than predicted.