2 replies to this topic

[movement K]

I'm working with primary mouse hepatocytes.
Currently I'm having a trouble with protein extraction from my cell culture.

I usually add Tris-HCl lysis buffer (supplemented with sucrose, KCl, MgCl2) and sonicate the samples.
However, when I spin the samples in the centrifuge, no matter how fast I spin them down, I get white stuffs (guessing genomic DNA) coagulated on top of the cell lysate.
This gives me a so much trouble for Bradford assay and loading samples on gels for Western Blotting.

Is there any ways to get rid of them?

Thank you

[vivekp]

movement K, on 07 February 2011 - 08:08 AM, said:

I'm working with primary mouse hepatocytes.
Currently I'm having a trouble with protein extraction from my cell culture.

I usually add Tris-HCl lysis buffer (supplemented with sucrose, KCl, MgCl2) and sonicate the samples.
However, when I spin the samples in the centrifuge, no matter how fast I spin them down, I get white stuffs (guessing genomic DNA) coagulated on top of the cell lysate.
This gives me a so much trouble for Bradford assay and loading samples on gels for Western Blotting.

Is there any ways to get rid of them?

Thank you

add some amount of Dnase....

[mdfenko]

i would think that the floating white material is lipid or carbohydrate.

Edited by mdfenko, 14 February 2011 - 02:11 PM.