Hoatlin:Western Protocol

BASIC WESTERN BLOT PROTOCOL

• Block membrane 1 hour in blocking buffer (5% carnation non-fat dry milk dissolved in water.)

• Incubate membrane for 1 hour in blocking buffer containing primary   antibody.

• Wash the membrane 2X5 min, 2X10 min in 0.05% Tween 20 in TBS.

• Incubate membrane for 30min in blocking buffer containing secondary antibody with desired (HRP, AP, etc.) conjugate.

• Wash the membrane 2X5 min, 2X10 min in 0.05% Tween 20 in TBS.

• Use ECL according to manufacturer to develop.

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