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RhoGTPase GST pull down assay

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#1 robby



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Posted 22 February 2009 - 07:02 AM

Hi all,

Is there anyone who measured the amount of active Rac using GST-PBD pull down assay? I have tried many times to measure Rac activity in cortical neurons. However the results were not consistent. I used 10 cm dish plated with 500 k cortical neuron and performed GST pull down after 5 days in vitro. The GST-PBD was purified from the E.coli and mobilized to beads with a concentration of 3 ug/ul. The protocol I used was from Upstate. Is there any tips for doing such an experiment? Thanks lot


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