Deglycosylation of ab108573. To examine deglycosylation of ab108573, 1µg of mouse progranulin (ab108573) was denatured with 1X Glycoprotein Denaturing Buffer at 100°C for 10 minutes. After the addition of NP-40 and G7 Reaction Buffer, two-fold dilutions of PNGase F were added and the reaction mix was incubated for 1 or 3 hour at 37°C. Separation of reaction products were visualized by immunoblotting with antibody against anti-mouse progranulin.
ERK phosphorylation induced by ab108573 in MCF10A cells. To examine the signal of phospho-p44/42 MAPK and p44/42 MAP kinase, reactions were carried out at 37°C over 0, 5, 10, 30, mins, respectively by adding ab108573 (500ng/ml) to the MCF10A human breast epithelial cells, which were maintained with serum starvation for 24hrs. Proteins in lanes 1, 2, 3, 4 were subjected to MCF10A human breast epithelial cell treatments over 0, 5, 10, 30mins, respectively.