pan-cadherin Antibody (CH-19): sc-59876

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	-Mouse (1:4000) -Use clean forceps to gently handle the blot from the corner without creasing the membrane. Do not write on the blot with pen or marker, as the ink can fluoresce and cause background.

Protocol tips
-Mouse (1:4000) -Use clean forceps to gently handle the blot from the corner without creasing the membrane. Do not write on the blot with pen or marker, as the ink can fluoresce and cause background.

Publication protocol

Dissected hippocampi were homogenized in protein lysis buffer (10mM Tris-Cl pH 6.8, 1.6 % SDS) containing protease and phosphatase inhibitor cocktails (Sigma). Supernatants were collected after centrifugation and the protein concentration was determined using a BCA assay kit (Thermo). Equal amounts of proteins (5 μg) were separated by electrophoresis on a 4 % – 12 % SDS–PAGE (Invitrogen), and then transferred to nitrocellulose membranes. After blocking with 5% BSA in TBS-T (Tris–buffer saline containing 0.1% Tween-20) for 1 hr at room temperature, membranes were hybridized with a primary antibody overnight at 4°C. After washing with TBS–T, membranes were incubated with a secondary antibody in 5% non-fat milk/TBS–T for 1 hr at room temperature. Signals were visualized by ECL (Thermo) and exposure time was adjusted so that the signals measured were in a linear range. After detecting phospho-Erk, the membranes were stripped and re-probed with a total Erk antibody. The total Erk levels were used to normalize each sample. The following primary antibodies were used: anti-phospho-Erk (#9101S, Cell Signaling, 1:6000), anti-total Erk (#9102S, Cell Signaling, 1:5000) and anti-SHP2 (sc-280, Santa Cruz, 1:3000).

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Manufacturer protocol

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