Cyclodextrin Screening Kit Data Sheet
Catalog No. 0602
The AthenaES™ Cyclodextrin Screening Kit is intended for identifi cation of the
best cyclodextrin for use in the refolding of proteins. Cyclodextrins’ unique
molecular structure greatly promotes protein refolding. Cyclodextrins can suppress
protein aggregation, protect against degradation, and alter the functions
of proteins. They also bind well to detergents (aggregation suppressors), which
allows for easy separation of the detergent from the protein, encouraging proper
refolding. The cyclodextrins remove the detergent from partially refolded protein
solutions, allowing the protein to complete the refolding process.
Different cyclodextrins act and respond to different protein structures. Since
no universal protein refolding condition has been identified, the best conditions
for a given protein must be determined empirically. Because of this, AthenaES™
provides 3 of the most commonly used cyclodextrins in a simple screening kit so
that the ideal conditions for the refolding of a given protein can be found quickly
and easily in a rapid screening format.
The Cyclodextrin Screening Kit can prove particularly useful when paired with
detergents and can be easily optimized with the AthenaES™ Detergent Screening
Kit. Individual cyclodextrins and detergents are also available.
||Store at Room Temperature.
DO NOT FREEZE.
||100 mM β-cyclodextrin
||100 mM methyl-β-cyclodextrin
||50 mM α-cyclodextrin
Instructions for Use
Cyclodextrins are typically used in a two-step protein refolding procedure. In
the first step the protein is partially refolded in the presence of detergent. It is
recommended that a suitable detergent be identified or that an experimental
design which screens both the detergent and the cyclodextrin simultaneously
be used. The following algorithm is suggested for identifying a suitable
Perform a preliminary screen of the physical-chemical parameters which are known
to affect protein refolding. Parameters to test include pH, ionic strength,
excipitents (i.e., detergents, polyols, chaotropic agents) redox state, temperature,
and protein concentration. Refolding can be done by dilution, dialysis, or
immobilization on a resin. Several commercial kits are available, such as Athena’s
QuickFold™ Protein Refolding Kit (Cat. No. 0600), which simplify the screening process by
providing the pre-mixed buffers along with straightforward statistical analyses of
the results. A detailed method can be found in the AthenaES™’s QuickFold™ Protein Refolding
Kit Application Manual at www.athenaes.com.
Identify a suitable detergent. Several different detergents should be tested to
determine the best one for the given target protein. Optimization of the detergent concentration
is not needed at this stage, but should be determined once the
cyclodextrin has been selected.
Perform the cyclodextrin screen. Perform the first step in the refolding process by
employing the buffer composition and detergent identified in steps 1 and 2. The
following protocol is recommended as it is one of the simplest to perform.
Prepare three reactions of the refolding buffer with the selected detergent and
add the denatured protein to each solution slowly while mixing gently to give a
final protein concentration of 50 µg/ml.
Incubate for 1 h on ice.
Add each cyclodextrin to a final concentration of 5 mM, one cyclodextrin per
reaction. Mix gently.
Incubate on ice for 1-24 h.
Assay the solution for refolded protein (by activity or physical means).
Once the cyclodextrin has been selected, the optimum concentration of the
detergent and cyclodextrin should be determined. The process can then be
adopted for an alternate refolding scheme such as by dialysis, diafiltration, or
Material Safety Data
FOR RESEARCH USE ONLY. NOT INTENDED OR APPROVED
FOR HUMAN, DIAGNOSTICS OR VETERINARY
USE. Do not ingest, swallow or inhale. Do not get in
eyes, on skin, or on clothing. Wash thoroughly after
handling. For complete safety information see full
Material Safety Data Sheet.