An alternative could be to disolve your cholesterol in methanol or maybe in 50% methanol, then add it directly to the drop. Does your protein tolerate 10-20% methanol? Then you can add the methanol-cholesterol solution you your hanging drops directly and mix fast. The methanol is volatile and will end up mostly in the reservoir leaving onlt traces in the drop, while the cholesterol will stay in the water. It might precipitate at first, but should redissolve or not precipitate at all if it binds your protein.
Jose Antonio Cuesta-Seijo. "Jerry McCully" <for-crystallizai...@hotmail.com> wrote: > > Dear ALL: > > Sorry for this kind of off-topic question. > > I am going to co-crystallize one protein with cholesterol. I read some > papers saying that > their protein can be pre-incubated with 1mM cholesterol in the presence of 5% > (v/v) ethanol. > > TO do so, I first dissolved cholesterol in 100% ethanol at a concentration of > 10mM. However, it is very difficult to make the dilution into 5% ethanol either > just in water or some buffers. > > Does anyone have such experience to make cholesterol solution in normal > buffers plus some ethanol? > > Thanks a lot, > > Jerry McCully > > _________________________________________________________________ > The New Busy think 9 to 5 is a cute idea. Combine multiple calendars with > Hotmail. > http://www.windowslive.com/campaign/thenewbusy?tile=multicalendar&ocid=PID28326::T:WLMTAGL:ON:WL:en-US:WM_HMP:042010_5 -- *************************** Jose Antonio Cuesta-Seijo Biophysical Chemistry Group Department of Chemistry University of Copenhagen Tlf: +45-35320261 Universitetsparken 5 DK-2100 Copenhagen, Denmark ***************************
