I have a crude extract from a cell suspension of bacteria that I broke with a French press. This extract is really dense and dark and for my enzymatic assays it causes issues due to the general concentration but mainly due to a pretty high viscosity. This viscosity can be seen after 3h ultracentrifugation at 180000g from an aliquot of my crude extract. After the ultracentrifugation we can see 2-3 layers having different colors and different viscosities while I expected something homogeneous. So, does anyone know how to keep the proteins in my substrate and remove a maximum of DNA and other stuff that I don't need? Also there is another concern, this must be done quickly because the enzymes targeted are oxygen sensitive. So, if someone has an idea to help me to deal with that, feel free to tell me.
One idea is to use CTAB ( Cetyl trimethyl ammonium bromide) to précipitate polyanions (DNA and polysaccharides). This will clarify the solution.
Hello professor, thanks for the answer, that's interest me but does it possible to have more details about how to use this CTAB: concentration, duration, before or after utracentrifugation, do I need to do another centrifugation if i add the CTAB after the first centrifugation? Thanks for your answer.
Well, you can try to add drop-wise a 1% CTAB solution to your crude extract until you cannot obtain more precipitate. But you have to check that the enzyme you search is still active.
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