The inhibitory effects of the 3E8 anti-HK antibody on Aβ42-induced HK cleavage are abolished by competitive HK-D6 peptides. (A) EDTA-human plasma was incubated with buffer, 3E8 anti-HK antibody (0.5 μM), control IgG (0.5 μM), HK-D6 (0.5 μM, 1 μM, 2 μM), 3E8 anti-HK antibody (0.5 μM) + HK-D6 (0.5 μM, 1 μM, 2 μM) or control IgG (0.5 μM) + HK-D6 (0.5 μM, 1 μM, 2 μM) (as indicated in [A]) at 37°C for 20 minutes. Aβ42 (5 μM) was added and incubated for 2 hours. Western blotting analyses were performed with antibodies against HK and TF. 3E8 HK antibody protected HK from Aβ42-induced cleavage (lane 3), but control IgG (lane 4) and different concentrations of HK-D6 did not have significant effects on Aβ42-induced HK cleavage (lanes 5-7 and [B]). However, HK-D6 dose-dependently inhibited the protective effects of 3E8 anti-HK antibody on Aβ42-induced HK cleavage (lanes 1, 3, 8-10, and [B]). HK-D6 did not have significant effects on the control IgG in Aβ42-induced HK cleavage (lanes 4, 11-13, and [B]). HK western blotting signals were normalized to TF. n = 6. Data are denoted as mean ± SEM. *P < .05, **P ≤ .01, ***P ≤ .001. P > .05 was not significant (n.s.).