Western Blot for weakly expressed protein - (Mar/25/2008 )
Hi, nice to meet you all here. Just a little question about Western Blot. In fact my protein is weakly expressed in the cell, is there any protocol or special kit for running a WB? Thanks.
Try immunoprecipitation and then western to detect. That'll enrich your protein.
you can amplify your detection by using a biotin-avidin (or streptavidin) system for your secondary antibody.
you have a biotinylated secondary and avidin (or streptavidin) and biotinylated detection enzyme (hrp or ap). this way you will have more than one enzyme molecule per antibody molecule.
in case your overexpressed protein carries a tag you may use it for specific enrichment before doing IB