Bio-Rad Rotofor® and Mini Rotofor Cells - Manual (Page 5)

Section 1
General Information
1.1 Introduction
Bio-Rad’s unique Rotofor System fractionates complex protein samples in free
solution using preparative isoelectric focusing. The Rotofor system is designed for
the initial clean up of crude samples and for use in purification schemes for the
elimination of specific contaminants from proteins of interest that might be difficult
to remove by other means.
The Rotofor cell provides up to 500-fold purification for a particular molecule in
less than 4 hours. Because electro-focusing is carried out in free solution, fractions
from an initial run can be easily collected, pooled and refractionated, resulting in up
to 1000-fold enrichment for a particular molecule. Purification using isoelectric
focusing is especially advantageous when protein activity needs to be maintained.
Bioactivity is maintained because the proteins remain in solution in their native
conformation.
The Rotofor cell incorporates a cylindrical focusing chamber with an internal
ceramic cooling finger. Rotation at 1 rpm around the focusing axis stabilizes
against convective and gravitational disturbances. Nineteen parallel, monofilament
polyester screens divide the focusing chamber into 20 compartments, each holding
one fraction. After focusing, the solution in each compartment is rapidly collected
without mixing using the harvesting apparatus supplied with the unit.
The Rotofor system is designed to accommodate a range of sample volumes
using interchangeable focusing chambers. The Mini Rotofor chamber is used for
sample volumes of 18 milliliters containing micrograms to milligrams of total protein.
The large Rotofor chamber is used for samples of 35 to 60 milliliters containing
milligrams to grams of total protein.
The Rotofor cell is used to purify a wide range of proteins. These include
monoclonal antibodies, cell surface receptor proteins, integral membrane proteins,
cytosolic and secreted enzymes, chemotactic factors, and recombinant proteins. It
has been used to separate isoenzymes, lipoproteins, and apolipoproteins.
Should a final purification step be required, we recommend the Model 491 Prep
Cell. The Prep Cell is a continuous elution gel electrophoresis device that uses
SDS-PAGE or Native-PAGE to completely purify individual proteins of interest. For
examples of published Rotofor cell applications, please refer to the Rotofor
Technical Folder (request Bulletin 1555A).
*Patent No. 4,588,492
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