His-Probe - (Aug/20/2013 )
i want to detect his-tagged protein in supernatant by using his-probe.
I applied supernatant supplemented with zeocin.
However, his-probe reacts with the supernatant containing zeocin alone...could this be true?
Are there histine groups in zeocin which led to a positive signal?
Is your his-probe an anti-his antibody or a nickel-based probe? Have you checked to see if the his-probe reacts with supernatant in the absence of zeocin (and your protein of course)? If your supernatant is from an insect cell line, they are known to have histidine rich proteins that could be confounding your results.
thank you, it seems that our nickel-based his-probe reacts with supernatant alone..