spun column

David L. Haviland, Ph.D. dhavilan at IMM2.IMM.UTH.TMC.EDU
Tue Aug 5 09:21:52 EST 1997


At 17:27 8/4/97 -0400, Molecular Pathology Lab wrote:
>Hello all-
>	I was wondering in anyone has a protocol for doing a G-50 spun column
>using a microcentrifuge for seperating unicorporated from labled probe. 
>I have used the 1mL (tuberculin) syringe method, but I don't have access
>to a clinical centrifuge, but I do have access to an old
>microcentrifuge.  Any help that you can give me will be greatly
>appreciated, and you can either send it to my e-mail or to the news
>group.

Eric:

Yes, we do all the time.  Here how... We use the Fischer Brand spin columns
#11-387-50 and we buy both G50 and G25 from Pharmacia.  The sepharose is
made per direction as resuspended final 50:50 with TE.  Our standard
columns are 3-4 mls.  We spin in a IEC but a JT6 or equivalent should
sufice.  A prep spin of 2 mins at 2000 rpm to remove excess water is done,
then the labeled sample is applied (tube swicth of course)  then spun again
4 mins at 2000 rpm.  Done.  Many times, I;ve prepped G25 with DEPC water
and used the column RNAse free.

Hope this helps,
David




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