Bio-Rad Prep Cell info
Casey Finnerty
jv...@galen.med.Virginia.EDU writes:
>Does anybody have experience with the apparatus sold by Bio-Rad to purify
>proteins? Our lab is in the process of trying one of them out.
>The unit's official designation is the "Model 491 Prep Cell" listing at
>about $2700 (US)
>We plan to use it to purify high molecular weight proteins in a
native-PAGE
>system.
>Any insights on the bugs inherent in the system? Efficacy of the system?
Our lab also tried this system out, with very poor results. Proteins
emerging from the gel end up being very dilute, provided the gel poured
is of sufficient height to result in decent separation. When we contacted
BioRad with this problem, they suggested pouring gels much shorter than
recommended in the manual. This resulted in less dilution but negligible
separation.
We were also interested in getting activity from the eluted proteins, but
the eluted protein was so dilute we didn't detect any. Admittedly, we
were using a relatively insensitive detection assay (hemagglutination).
Also, in order to maximize recovery of activity, we tried the Jovin
buffer system that keeps the pH closer to neutral than the standard
Tris-glycine system. The buffer components of the Jovin system are not
cheap, and the 491 Prep cell consumes a lot of buffer.
I you are still interested in trying preparative electrophoresis, a
homemade system is described by Shain et al in Anal. Biochem. 200, 47-51,
1992. Also there is an article by Ugozzoli and Chin in Biotechniques
(12(12), 187-188, 190) on preparative agarose electrophoresis of large MW
proteins.
If you decide to buy a BioRad separation system, I would recommend
instead the Rotofor system. If you can prevent protein precipitation and
don't overload the cell, the separations are usually effective and very
simple to perform.
Casey M. Finnerty
Boyce Thompson Institute at Cornell University
cm...@cornell.edu
607-254-1262