94
AFC
TABLe I
Characteristics of TSKgel AFC columns
INTRODUCTION TO TSKgel AFFINITY CHROMATOGRAPHY COLUMNS
The TosohBioscience TSKgel Affinity Chromatography (AFC) column line
consists of two group-specific stationary phases:
TSKgel Boronate-
5PW
and
TSKgel Chelate-5PW
as well as one activated packing
material called
TSKgel Tresyl-5PW
. Affinity chromatography offers the
highest level of specificity and selectivity in biomolecular separations
and purifications. Tosoh Bioscience supplies a full range of products
for analytical, preparative and process scale affinity chromatography.
TSKgel affinity chromatography columns are based on the well-known
G5000PW porous resin, which is the basis for high performance size
exclusion chromatography columns. The TSKgel 5PW-type resin is a
hydrophilic media with 100 nm pores and an estimated protein exclusion
limit of 5 x 10
6
Da. Tosoh Bioscience’s process scale affinity media are
based on the 65 µm particle size, semi-rigid TOYOPEARL HW-65 resin.
Since analytical and semi-preparative columns are made from the same
polymer chemistry as the process scale media, seamless scale-up from
lab to process scale is achievable. Consult the chapter on bulk media
for more information about resins for packing columns to purify medium
to large volume samples.
Column Selection
Table I
lists the ligand concentration, adsorption capacity and the test
analyte used to determine the capacity of each column type.
The structures of the functional ligands available from Tosoh Bioscience
are shown in
Figure 1
. The choice of a specific ligand is dictated by
the expected interaction between the sample and column bonded
phase. For example, the TSKgel Chelate-5PW column will bind high
concentrations of Zn
2+
ions. If a given protein is known to bind to Zn
2+
ions, the Chelate-5PW would be a candidate column for the isolation of
that target compound.
Tosoh Bioscience offers AFC columns in both glass and stainless steel
formats. Glass columns are available in 5 mm ID x 5 cm L and 8 mm ID
x 7.5 cm L. Stainless steel columns are available as 7.5 mm ID x 7.5 cm
L and 6 mm ID x 4 cm L (Tresyl-5PW only). TSKgel BioAssist Chelate
is packed in 7.8 mm ID x 5 cm L PEEK hardware. The shipping solvent
is distilled water for Boronate-5PW. The Chelate-5PW is shipped in 10
mmol/L acetate buffer, pH 4.5, and the Tresyl-5PW column shipping
solvent is acetone.
Stainless steel or Pyrex frits are employed in the body of the column end-
fittings for themetal and glass columns, respectively. The nominal frit size
forstainlesssteelcolumnsisengravedintheend-fittingsandallPyrex
®
frits
are 10 µm nominal pore size.
FEATURES
BENEFITS
High size exclusion limit (> 5 x 10
6
Da)
Enhanced access of large proteins to affinity ligands
Small particle size
High efficiency for analytical (10 µm) and semi-preparative (13 µm)
affinity applications.
Rigid polymeric base resin
Wide pH range (2-12) of the base resin, enabling robust cleaning
options
Stable affinity ligands
Long lifetime, solvent compatibility, autoclavable
Choice of four affinity ligands
Application flexibility, scalability from lab to commercial
production.
TSKgel BioAssist Chelate offered in PEEK hardware
Eliminates undesirable interactions with column hardware.
Column packing
Ligand type
Ligand concentration
Adsorption capacity
Sample
Boronate-5PW
m-aminophenyl-boronate
not available
40 µmol/mL resin
sorbitol
Chelate-5PW
iminodiacetic acid
20 µmol/mL resin
not available
not available
Tresyl-5PW
tresyl
ca. 20 µmol/mL resin
>60 mg/g dry resin
(coupling capacity)
soybean trypsin inhibitor
