I just meant that you may have had to keep the sample at 37C to keep the phases separate during centrifugation, but that's assuming they even existed in the first place. I'm only guessing, since I've never considered running the protocol with alcohol in the solution - it probably did just keep the Triton soluble the whole time.
Oh and regarding Nathan's post, the Triton+endotoxin will definitely be in the bottom layer, not the top.
On Wednesday, February 6, 2013 3:32:18 PM UTC-8, phillyj wrote:
-- On Wednesday, February 6, 2013 3:32:18 PM UTC-8, phillyj wrote:
On Wed, Feb 6, 2013 at 6:18 PM, Max Berry <maxb...@gmail.com> wrote:
> Yeah the 15% isopropanol is definitely gonna keep the triton soluble so it
> wouldn't separate - I forgot Qiagen uses isopropanol in those buffers. If
> you do resuspend in water, the phase separation protocol should work in
> that, or you can add a little triton into the load before you put in back on
> the column, which might help on the next run. Actually, it might well have
> separated at 37C even with the isopropanol, but if your centrifuge isn't
> heated then it might have cooled down and resolubilized again.
>
I see. Thanks for the input. I don't understand what you mean about
the heated centrifuge? I centrifuged at room temperature (25-27C) so
you're saying that it resolubilized then? I didn't really notice any
phase after taking it out from the 37C but maybe I wasn't paying
enough attention.
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