FIGURE 8:

The p10 N-terminal domain of p35 preferentially binds multimeric proteins in cortical neurons transfected with p35 myc-his. Rat cortical neurons at 5 days in vitro were transfected with p35 myc-his or p25 myc-his plasmid. After 48 h, lysates were prepared and subjected to SDS–PAGE using 30 μg of lysate for each lane. (A) Protein expression was analyzed using three different antibodies to confirm p35 and p25 expression with p35 (C-19), myc, and his antibodies, respectively. Arrowhead and arrow represent the expression of myc-his p35 and myc-his p25, respectively. (B–F). Both p35 and p25 lysates were used at 500-μg protein concentration for immunoprecipitation with mouse monoclonal myc antibody. Input represents expression level of each protein. After SDS–PAGE, Western blotting was performed using different polyclonal antibodies as follows: p67 (B), PSD95 (C), tau (D), GFAP (E), and syntaxin 1 (F). Data were analyzed with Prism 5.0 software. Tukey’s multiple comparisons test was used. Differences with *p < 0.05 were considered significant. The quantification of bands was done using ImageJ software. The results represent the value of three experiments and are expressed as ±SEM. **p < 0.01 and ***p < 0.001.