Hi all,
Is it possible to redevelop an EMSA blot which has already been exposed? I am using ThermoScientific LightShift Chemiluminescence kit and believe that my blot could be improved if I had left the developing reagent longer before exposing the blot. Is there a way to go back and re-do it, without performing the experiment again?
Thanks
Hi, I am not sure about exactly EMSA blot, for general blots I can suggest. If you don't have enough signal then you can reincubate it for longer time and expose for longer time. We first use ECL from millipore if the signal is weak we use famto ECL from thermo scientific. You don't need to re-blot just to re-develope (may be just include a washing step). If the signal is bleached you can try to wash the membrane well and re-do your secondary antibody and then develop. Hope it helps.
Yes, you can do one-time reproducibility in EMSA, wash blot one or two times, then keep it in 4 degrees C (only membrane using for next day), restart your experiment starting from blocking steps, and followed by HRP-streptavidin incubation. (Note imp step: oligo in the nitrocellulose blot is fixed well only if it exposing to UV light).
Good luck
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