I generally use 1 - 2% DDM for extraction only, but lower the concentration to 0.01% for following steps (i.e. NiNTA and gel filtration). The excess DDM is washed away by using a lower concentration in your wash and elution buffers.
Kelly ******************************************************* Kelly Daughtry, Ph.D. Post-Doctoral Fellow, Raetz Lab Biochemistry Department Duke University Alex H. Sands, Jr. Building 303 Research Drive RM 250 Durham, NC 27710 P: 919-684-5178 ******************************************************* On Mon, Mar 26, 2012 at 1:17 PM, Katarzyna Rudzka <kasiarud...@yahoo.com>wrote: > Hi All, > Has anyone had any luck purifying membrane proteins with DDM > (n-dodecyl-D-maltoside) using concentration of detergent ~1-2 x CMC ? (Its > CMC is very low: 0.009%). I would like to keep it as low as possible, so I > don't have too much DDM around when I get to the crystallization step. I > wonder If the amount of detergent sufficient for the protein extraction has > to be determined experimentally for each protein or maybe there are some > good rules of thumb. I appreciate your help. Thanks. > Kasia > > > Katarzyna Rudzka, Postdoctoral Fellow > Department of Biophysics and Biophysical Chemistry > Johns Hopkins University, School of Medicine > Baltimore, Maryland 21205 USA >
