Phospho-JNK, Santa Cruz Biotechnology, sc-6254

This antibody was tested on 50 ug lysate from mouse primary hippocampal neurons.
The antibody was tested simultaneously as p-JNK from Biosource #44-682G (see full review here ( http://pabmabs.com/wordpress/?p=874 ). While an excellent result was obtained using 44-682G, this antibody did not work at all. Many (intense) unspecific bands were observed, and none of these corresponded to the correct size. the buffer used to lyse the cells is shown below.

• 20 mM MOPS, pH 7.0 (any other buffer at this pH could be substituted);
• 2 mM EGTA (to bind calcium);
• 5 mM EDTA (to bind magnesium and manganese);
• 30 mM sodium fluoride (to inhibit protein-serine phosphatases);
• 60 mM β-glycerophosphate, pH 7.2 (to inhibit protein-serine phosphatases);
• 20 mM sodium pyrophosphate (to inhibit protein-serine phosphatases);
• 1 mM sodium orthovanadate (to inhibit protein-tyrosine phosphatases);
• 1% Triton X-100 (can be substituted with 1% Nonidet P-40) Important Note: Do not add if you intend to first prepare a cytosolic fraction.
• 1 mM phenylmethylsulfonylfluoride (to inhibit proteases);
• 3 mM benzamidine (to inhibit proteases);
• 5 µM pepstatin A (to inhibit proteases);
• 10 µM leupeptin (to inhibit proteases);
• 1 mM dithiothreitol (to reduce disulphide linkages)

adjust to pH 7.2