ResourceLife Sciences
Validating the Expression Consistency of a Housekeeping Protein
16 Jun 2013Housekeeping proteins (HKP) are typically used as loading controls because it is assumed that the HKP expression level remains consistent across samples. However, there is evidence that HKP expression levels change in many scenarios, including siRNA treatment, cell death, cell differentiation, etc. Therefore, validation of consistent HKP levels under specific experimental conditions is suggested prior to its selection as a loading control. This application note describes how to validate the consistency of HKP expression before the actual western blot experiment is performed.
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Western BlottingWestern blotting equipment is used to transfer and identify specific proteins within a sample, reveal protein modifications, as well as give a semi-quantitative estimation of their concentration. Western blotting equipment includes all apparatus necessary to transfer proteins from gel to membrane and subsequent processing steps. Protein transfer can be performed by electroblotting with wet, semi-dry and dry transfer systems onto nitrocellulose and PVDF membranes. Blocking, washing and labeling of membranes follows, involving buffers, blocking reagents, blotting / incubation trays, labeling reagents, immunoblotting assays, antibodies and conjugates. Automated equipment for these steps is available to accelerate your lab workflow. Finally, detection and imaging of proteins can be conducted using gel documentation and imaging systems. Find the best western blotting equipment in our peer-reviewed product directory: compare products, check customer reviews and receive pricing direct from manufacturers.ValidationHousekeeping ProteinLoading ControlProtein Quantitation