Figure 6
From: Biparatopic anti-PCSK9 antibody enhances the LDL-uptake in HepG2 cells

Western blotting and quantitative analyses. The western blotting and quantitative analyses were performed to determine the expression level of LDLR (~ 150 kDa) of hepatocytes. The α-tubulin (~ 55 kDa) was served as the internal control protein. Because the LDLR and α-tubulin are incubated with different Abs, we did not perform co-incubation to avoid non-specific bands. Under the same conditions, each sample was divided equally into two equal parts. Then they were loaded to two same SDS-PAGE gels, one for the test of LDLR and the other for α-tubulin. The full-length blots were shown in Fig. S4A-D. The results are presented in the form of high (1.5 μM) (A), medium (0.75 μM) (C) and low (0.375 μM) (E) doses. BC: the blank control group, referring to no addition of PCSK9 and its inhibitor; NC: the negative control, referring to only addition of 0.1 μM PCSK9. “+” and “−” refers to the addition and absence of the reagents such as LDL-BODIPY, PCSK9 or its Abs in above LDL-uptake assays. And “anti-LDLR-Ab, +” refers to the addition in the western blotting assays. The quantitative analyses of high (1.5 μM) (B), medium (0.75 μM) (D) and low (0.375 μM) (F) doses were performed by software “ImageJ”. The blotting results shown are representative of at least two independent experiments. Normalization was processed by dividing the greyscale value of the experimental group by that of the respective α-tubulin protein. Statistical significance was determined using Student’ s paired t-test. P < 0.05 was considered as statistically significant, compared with the negative control group (*P < 0.05, **P < 0.005, ***P < 0.0005, ns: not significant).