Monoclonal mouse anti pan cadherin

Membrane-enriched whole-brain or hippocampal tissue homogenates (15–30 ug) were subjected to SDS-PAGE electrophoresis as previously described (Makinson et al., 2014 (link)). After blocking in 5% milk, blots were incubated overnight at 4°C in either polyclonal rabbit anti-Na_v1.6 primary antibody (1:200, Millipore, Billerica, MA), polyclonal rabbit anti-Na_v1.6 (1:225, Alomone, Israel), polyclonal rabbit anti-Na_v1.1 (1:200, Millipore), or monoclonal mouse anti-Na_v1.2 (1:1000, Neuromab, Davis, CA). Blots were then incubated in either HRP-conjugated goat anti-rabbit secondary (1:10,000, GE Healthcare, United Kingdom), HRP-conjugated goat anti-mouse secondary (1:10,000, Jackson ImmunoResearch, West Grove, PA), or HRP-conjugated goat anti-rabbit secondary (Sigma, St. Louis, MO, 1:16,000) for 1 hr followed by washing in SuperSignal West Pico Chemiluminescent substrate (Thermo Fisher) and imaging. Blots were also probed using a monoclonal mouse anti-α-tubulin (1:10,000, Millipore) or monoclonal mouse anti-pan-cadherin (1:100,000, Sigma) antibody followed by HRP-conjugated goat anti-mouse secondary (1:10,000, Jackson ImmunoResearch) or HRP-conjugated goat anti-mouse secondary (Pierce, 1:26,000) for normalization of sample loading. Image quantification was performed using ImageJ software (NIH).

Membrane-enriched whole-brain or hippocampal tissue homogenates (15-30 ug) were subjected to SDS-PAGE electrophoresis. After blocking in 5% milk, blots were incubated overnight at 4°C in either polyclonal rabbit anti-Na_v1.6 primary antibody (1:200, Millipore, Billerica, MA), polyclonal rabbit anti-Na_v1.6 (1:225, Alomone, Israel), polyclonal rabbit anti-Na_v1.1 (1:200, Millipore), or monoclonal mouse anti-Na_v1.2 (1:1000, Neuromab, Davis, CA). Blots were then incubated in either HRP-conjugated donkey anti-rabbit secondary (1:10,000, GE Healthcare, United Kingdom), HRP-conjugated goat anti-mouse secondary (1:10,000, Jackson ImmunoResearch, West Grove, PA), or HRP-conjugated goat anti-rabbit secondary (Sigma, St. Louis, MO, 1:16,000) for 1 h followed by washing in SuperSignal West Pico Chemiluminescent substrate (Pierce) and imaging. Blots were also probed using a monoclonal mouse anti-α-tubulin (1:10,000, Millipore) or monoclonal mouse anti-pan-cadherin (1:100,000, Sigma) antibody followed by HRP-conjugated goat anti-mouse secondary (1:10,000, Jackson ImmunoResearch) or HRP-conjugated goat anti-mouse secondary (Pierce, 1:26,000) for normalization of sample loading. Image quantification was performed using ImageJ software (NIH).