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Western blotting is the technique used to identify and locate proteins based on the ability to bind to specific antibodies cells and tissues need to be lies to release the proteins of interest preparation of the lies aid from the tissues is done by addition of phosphate buffer centrifuge 20 minutes at 12,000 rpm at 4 degree Celsius in a micro centrifuge remove the tubes from the centrifuge in place on eyes discard the pellet determine the protein concentration by Bradford or Lori assay electrophoresis can be one-dimensional or two-dimensional sds-page technique is a standard means for separating proteins ing to their molecular weight the separation of molecules within a gel is determined by the relative size of the pores formed within the gel the proteins in the tested solution are separated into distinct bands by sds-page the gel should be soaked in transfer buffer for 10 minutes a nitrocellulose membrane approximately the size of the gel must be pre soaked in Western transfer buffer
