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| Comments : The antibody is the best phospho-IKK Ser 176/180 ab I have ever used. I stimulated insolated mammary gland tumor cells with LPS at 1 ug/ml. The antibody gave out every a specific band which merged with the upshifted form of IKKbeta in the IKKbeta rebloting.
I blot the membrane with 5% milk in washing buffer. I did 1:200 dilution in 2.5% milk in washing buffer for the primary antibody and incubate the membrane overnight at 4 ºC. After washes, I apply 2nd ab (pierce) at 1:2000 dilution in washing buffer. After washes, the membrance is developed with pierce long duration ELC kit.
NOTE: washing buffer 0.1% Tween20 and 10mM NaF in PBS)
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