2.14 Immunoprecipitation assay
After the treatment of Lf and/or SB203580 for 24 h, the N2a-sw cells
were solubilized in NP40 buffer supplemented with protease inhibitor and
PMSF, one part of the protein extracts were used as the input control,
and the rest (~1 mg) was incubated with rabbit
anti-PP2Ac antibody (Cell Signaling Technology, 2038) (another extract
from untreated N2a-sw cells was incubated with IgG as the negative
control) overnight at 4 °C, and followed by precipitation with protein
A-beads (Thermo Fisher Scientific, 10001D) for 4 h at 4 °C. The beads
were washed and added with 50 μl sample buffer, the mixture was boiled
for 5 min and sedimented, and the supernatant was used for western blot
analysis.