APP-BP1, PS1 and nicastrin co-immunoprecipitated and co-migrated in brain protein extracts. A. APP-BP1 co-immunoprecipitated with PS1 in brain lysates. Adult rat brain lysates were immunoprecipitated with the rabbit anti-PS1 antibody (middle lane), or a non-related antibody (rabbit anti-cyclin B1). The blot was probed with the anti-APP-BP1 antibody, BP339 or with rabbit anti-nicastrin. Anti-PS1 specifically precipitated APP-BP1 and nicastrin which were absent in control lanes. This experiment has been repeated three times. B. PS1 co-precipitated with APP-BP1 in membrane fractions. Proteins extracted from membrane fractions were precipitated with the APP-BP1 antibody, BP339 or with the BP339 preimmune serum (pre) or beads alone. The blot was probed with the PS1 antibody (AB5308) for PS1-FL and PS1-CTF, or with antibodies against PS1-NTF or nicastrin. C. APP-BP1 co-migrated with PS1 in glycerol gradient of membrane protein extracts isolated from adult rat brains. Brain membrane proteins were subjected to glycerol gradient centrifugation. Proteins in equal volumes from each fraction were resolved by SDS-PAGE gels. The blots were incubated with specific antibodies followed by ChemiGlow detections. Fractions 2 to 4 contain the majority of each protein analyzed. APP-BP1 was found to co-migrate with PS1 (FL, NTF, and CTF), and nicastrin. The location of β-catenin was also shown in the glycerol gradient. BioRad Precision Plus protein standards are shown on the right. PS1-CTF identified by AB5308 antibody was verified by a PS1-C-specific antibody (D).