Figure 2.

Immunoblotting with BACE-Cat1 reveals elevated BACE1 levels in human AD and APP transgenic mouse brains. A, Human autopsy brain tissues (temporal lobe cortex) of clinically diagnosed AD (n = 3) and non-demented controls (ND) (n = 3) were homogenized and analyzed by BACE-Cat1 immunoblot. Lanes with BACE1−/− (KO) brain homogenate and lysate from BACE1-overexpressing HEK293 cells (+) were included as negative and positive controls, respectively. Molecular mass markers are on the left. Immunoblot analysis for β-actin served as a loading control. B, Brain homogenates from representative 2- to 12-month-old 5XFAD (n = 3–5 per age) and age-matched normal control (Non-Tg) (n = 3–5 per age) mice were subjected to immunoblot analysis using BACE-Cat1 antibody. The BACE1 band migrates at ∼70 kDa, and the occasional band at ∼60 kDa represents endogenous mouse IgG (ms IgG) (detected by goat anti-mouse IgG secondary antibody) present in blood of brains from mice that were not transcardially perfused with buffer. Note that the lane with the homogenate from perfused KO brain does not have the background band. The intensities of the BACE1 bands in 5XFAD lanes are significantly greater than those of age-matched Non-Tg BACE1 bands. C, Quantification of 5XFAD and Non-Tg BACE1 signals detected by BACE-Cat1 immunoblot analysis is represented as percentage of 2 month Non-Tg control (100%). Note that BACE1 levels in 5XFAD brains reach ∼190% of control at 9 months and level off and that Non-Tg BACE1 levels appear to decrease slightly with age. Error bars indicate SEM. ***p < 0.001.