LC COLUMNS Tosoh Bioscience - Hichrom€¦ · HICHROM Chromatography Columns and Supplies Hichrom...

HICHROMChromatography Columns and Supplies

Hichrom Limited1 The Markham Centre, Station RoadTheale, Reading, Berks, RG7 4PE, UK

Tel: +44 (0)118 930 3660 Fax: +44 (0)118 932 3484Email: [email protected] www.hichrom.co.uk

Catalogue 9

LC COLUMNSTosoh Bioscience

LC Columns – Tosoh Bioscience

www.hichrom.co.uk • [email protected] • Tel: 0118 930 3660 243

Hichrom Lim

itedTosoh Bioscience

TOSOH BIOSCIENCE

Tosoh Bioscience is a major supplier of liquid chromatography products in the pharmaceutical and biotechnology industries, for the analysis, isolation and purification of proteins, peptides, oligonucleotides and enzymes as well as low molecular weight compounds. TSKgel® is the most popular of several ranges of columns and bulk packing materials supplied by Tosoh Bioscience. An extensive range of products is offered for size exclusion, ion-exchange, reversed-phase, HILIC, HIC and affinity separation techniques. Both silica-based and polymer-based materials are available.

TSKgel columns are available for both modes of size exclusion chromatography (SEC) – gel filtration chromatography (GFC) of water soluble polymers in aqueous eluents and gel permeation chromatography (GPC) of organic soluble polymers using non-aqueous eluents. The TSKgel SW and PW column lines are suitable for GFC, the H-type columns for GPC and the Alpha and SuperAW Series can be used for both GFC and GPC.

Silica-based Phases (SW)• Rigid spherical porous silica• Hydrophilic bonding (diol groups)• Stable from pH 2.5 – 7.5• Widely referenced• Three different pore sizes

TSKgel SW size exclusion columns are widely used for aqueous separation of proteins, antibodies, enzymes, nucleic acids and other biological macromolecules. Columns show high resolution and sensitivity and have excellent solvent stability with up to 100% polar organic solvents. The smaller particle size of the SWXL materials results in reduced analysis times. All SW-type columns are available in stainless steel hardware; SWXL columns are also available in PEEK hardware. TSKgel G3000SWXL is the ideal scouting column for unknown molecular weight compounds. If the protein of interest elutes near the exclusion volume, then G4000SWXL is the next step. Conversely, if the protein of interest elutes near the end of the chromatogram, then G2000SWXL should be considered.

The Super SW2000 and SW3000 materials (4μm) provide fast, high efficiency separations. The increased sensitivity compared with the corresponding SWXL or SW phase is especially useful for limited sample quantities. Figure 1 shows the separation of a standard protein mixture on TSKgel SuperSW3000.

TSKgel SW Phase

Particle Size (μm)

Pore Size (Å)

Exclusion Limit (Da) for globular proteins

SuperSW2000 4 125 5,000 – 150,000

G2000SWXL 5 125 5,000 – 150,000

G2000SW 10, 13 125 5,000 – 150,000

SuperSW3000 4 250 10,000 – 500,000

G3000SWXL 5 250 10,000 – 500,000

G3000SW 10, 13 250 10,000 – 500,000

G4000SWXL 8 450 20,000 – 7,000,000

G4000SW 13, 17 450 20,000 – 7,000,000

TSKgel® Silica SEC Phases

Ordering Information – Silica SEC Phases

TSKgel SW Phase

Column Dimensions (mm) Guard Column

Analytical Preparative

Analytical Preparative 300 x 4.6 300 x 7.5 600 x 7.5 300 x 7.8 300 x 21.5 600 x 21.5

£1,635 £1,380 £2,087 £1,438 £3,554 £5,170

SuperSW2000 18674 - - - - - 187621 £455

-SuperSW3000 18675 (£1,578) - - - - -

G2000SW (10μm) - 5788 5102 - 6727 (13μm) 5146 (13μm)53712 £397

57584 £961

G3000SW - 5789 5103 - 6728 (13μm) 5147 (13μm)

G4000SW (13μm) - 5790 5104 - 6729 (17μm) 5148 (17μm)

G2000SWXL - - - 8540 - -

85433 £397

- G3000SWXL - - - 8541 - -

- - - £1,488 - -

G4000SWXL - - - 8542 - -1 Dimensions 35 x 4.6mm 2 10μm. Dimensions 75 x 7.5mm 3 Dimensions 40 x 6.0mm 4 Dimensions 75 x 21.5mm

Figure 1. Standard protein mixture on TSKgel SuperSW3000

1. Thyroglobulin2. Bovine IgG3. Ovalbumin4. Myoglobulin5. Cyanocobalamin

Column dimensions: 300 x 4.6mmEluent: 0.1M Na phosphate, pH 6.7 – 0.2M Na sulphateFlow rate: 1ml/minTemperature: ambient

Size Exclusion Chromatography Phases

0 10 20 Mins

1

2

4

3

5

Please contact Hichrom for current prices. Tel: +44 (0)118 930 3660 Email: [email protected]

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Size Exclusion Chromatography Phases (continued)

Polymer-based Phases (PW)• Hydrophilic, rigid spherical porous methacrylate beads• Excellent chemical and mechanical stability• Stable from pH 2 – 12• Wide molecular weight range• Temperature stable up to 80°C

Polymeric TSKgel® PW and PWXL columns are designed for GFC of water soluble organic polymers, polysaccharides, oligosaccharides, DNA and RNA. They are based on a hydrophilic polymethacrylate matrix, which is stable from pH 2 to 12 and in aqueous eluents with up to 50% polar organic solvent.

The TSKgel PWXL range is preferred for analytical purposes, due to increased efficiency and resolution. Figure 2 shows typical polyethylene glycol and oxide calibration curves obtained with TSKgel PWXL columns. For preparative work, the 600mm length TSKgel PW columns are recommended due to their increased loading capacity. The mixed bed TSKgel GMPW and GMPWXL columns are useful for samples with a broad molecular weight range.

TSKgel PWXL-CP columns are especially suited for the separation of cationic polymers (see below).

TSKgel® Polymeric SEC Phases

TSKgel PW PhaseParticle Size

(μm)Pore Size

(Å)

Exclusion Limit (Da)

Polyethylene glycols and oxides

Dextrans

G1000PW 12 <100 < 1,000 -

G2000PW 12 125 < 2,000 -

G2500PW 12, 17 <200 < 3,000 < 3,000

G3000PW 12, 17 200 < 50,000 -

G4000PW 17 500 < 300,000 -

G5000PW 17 1,000 < 1,000,000 -

G6000PW 17 >1,000 < 8,000,000 -

GMPW 17 <100-1,000 500 – 8,000,000 -

G2500PWXL 7 <200 < 3,000 -

G3000PWXL 7 200 < 50,000 < 60,000

G4000PWXL 10 500 < 30,000 1,000 – 700,000

G5000PWXL 10 1,000 < 1,000,000 50,000 – 2,500,000

G6000PWXL 13 >1,000 < 8,000,000 500,000 – 50,000,000

GMPWXL 13 10-1,000 500 – 8,000,000 < 50,000,000

G3000PWXL-CP 7 200 < 90,000 -

G5000PWXL-CP 10 1,000 < 1,000,000 -

G6000PWXL-CP 13 >1,000 < 20,000,000 -

SuperMultipore PW-N 4 N/A 300 – 50,000 -

SuperMultipore PW-M 5 N/A 500 – 1,000,000 -

SuperMultipore PW-H 8 (6-10) N/A 1,000 – 10,000,000 -

TSKgel PWXL-CP SeriesTSKgel PWXL-CP columns are designed to facilitate the separation of water soluble cationic polymers by SEC. They are produced in three different pore sizes for different molecular weight ranges. By incorporating a cationic functionality on the polymethacrylate particle surface, ionic adsorption of the polymers is eliminated. This modification results in high recovery for ationic polymers and enables elution under low salt conditions.

Figure 3 shows successive injections (from red to black traces) of polyallylamine. Similar elution profiles were obtained with no loss of polymer due to adsorption.

-20

20

60

100

140

180

3 5 7 9 11 13

Elution time (min)

mV

Column: TSKgel G5000PWXL

-CPEluent: 0.1mol/L NaNO

3

Flow rate: 1.0mL/minDetection: RITemperature: 25°CSample: polyallylamine-HCl (PAA)Sample load: 3g/L, 100µL

TSKgel PWXL columns (300 x 7.8mm)H: G2500PWXL J. G3000PWXL

K: G4000PWXL L: G5000PWXL M: G6000PWXL N: GMPWXL

Eluent: Distilled H2O

Flow rate: 1.0ml/min

Figure 2. TSKgel PWXL columns

5 10 15Elution volume (mL)

Figure 3. Elution profiles of PAA on TSKgel G5000PWXL-CP

Column: TSKgel G5000PWXL-CP (300 x 7.8mm)Eluent: 0.1M NaNO

3 Flow rate: 1ml/min

Temperature: 25°C Detection: RISample: Polyallylamine. HCl (PAA)

3 5 7 9 11 13

Elution time (mins)

102

103

104

105

106

MW



Hichrom Lim



Polymer-based Phases (PW) (continued)

TSKgel® SuperMultiporePW SeriesTSKgel® SuperMultiporePW columns are packed with monodisperse polymethacrylate particles, each containing a wide range of pore sizes to cover different molecular weight ranges (see Figure 4). Multi-pore particle technology is the best way to achieve near linear SEC calibration curves, without the peak disturbances which typically occur due to a mismatch of pore sizes from coupled columns with different molecular weight ranges. This is illustrated in Figure 5 for the analysis of polyvinylpyrrolidone.

Large Pore Medium Pore Small Pore Multiple Pore Size

Pure packingswith multi-pore size distribution

Blend (mixed bed) packingsof different grades

Connect columns withdifferent grades of packings

Conventional Strategy New Strategy

Strategies for wide range separation using Size Exclusion Chromatography

Ordering Information – Polymeric SEC Phases

TSKgel PW PhaseColumn Dimensions (mm) Guard Column

Analytical Preparative

Analytical Preparative150 x 6.0 300 x 7.5 300 x 7.8 600 x 7.5 600 x 21.5

- £1,388 - £1,949 -

G1000PW - 5760 - - -67631 £419 -

G2000PW - 5761 - 5105 -

G3000PW - 5762 - 5106 -

67621 £419 67585 £1,008

G4000PW - 5763 - 5107 -

G5000PW - 5764 - 5108 -

G6000PW - 5765 - 5109 -

- £1,196 £1,334 £1,949 £4,918

G2500PW - 8028 - 8029 8030 (17μm)

GMPW - 8026 - 8027 -

G2500PWXL - - 8020 - -

80332 £397

-

G3000PWXL - - 8021 - -

G4000PWXL - - 8022 - -

G5000PWXL - - 8023 - -

G6000PWXL - - 8024 (£1,309) - -

GMPWXL - - 8025 - -

£1,377 - £1,436 - - -

G3000PWXL-CP - - 21873 - -

218763 £429G5000PWXL-CP - - 21874 - -

G6000PWXL-CP - - 21875 - -

SuperMultipore PW-N 22789 - - - - 227934 £417

SuperMultipore PW-M 22790 - - - - 227944 £417

SuperMultipore PW-H 22791 - - - - 227954 £4171 13μm. Dimensions 75 x 7.5mm 2 12μm. Dimensions 40 x 6.0mm 3 13μm. Dimensions 40 x 6.0mm 4 Dimensions 35 x 4.6mm 5 17μm. Dimensions 40 x 6.0mm

Figure 4. Strategy for wide range separation using SEC

Columns: TSKgel SuperMultiporePW-M (150 x 6.0mm) (red) TSKgel G3000PWXL and G5000PWXL (each 300 x 7.8mm) in series (blue)Eluent: 0.1M NaNO

3

Flow rate: 0.6ml/min

Figure 5. Analysis of polyvinylpyrrolidone

1.5 3.5 5.5 7.5 9.5 11.5Elution time (mins)


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Polymer-based Phases (Alpha and SuperAW)• Unique hydrophilic polyvinyl resin• Wide solvent compatibility, from 100% water to 100% non-polar organic solvents• Strong mechanical stability

The TSKgel® Alpha and SuperAW series columns are based on a hydrophilic, highly cross-linked vinyl polymer resin. These materials are mechanically stable and exhibit minimal swelling and shrinkage. They are compatible with solvents from pure aqueous to 100% organic (except chloroform, toluene and hexane), making them suitable for both GFC and GPC. They have applications in the analysis of polar synthetic oligomers, polysaccharides and detergents. The TSKgel Alpha series consists of 6 columns with different pore sizes and 3 particle sizes for a wide molecular weight separation range. The TSKgel SuperAW series offers the benefit of smaller particle sizes and smaller column dimensions, making them ideal for high throughput applications. For samples with big differences in molecular weights, the mixed bed columns TSKgel Alpha-M and TSKgel SuperAWM-H show linear calibration curves over the whole range.

Ordering Information – TSKgel® Alpha and SuperAW PhasesTSKgel Phase Particle Size (μm) Exclusion Limit (PEO/H2O) Column Dimensions (mm) Cat. No. Price

Alpha-25001 7 5,000 300 x 7.8 18339 £1,217

Alpha-30001 7 90,000 300 x 7.8 18340 £1,217

Alpha-40001 10 400,000 300 x 7.8 18341 £1,217

Alpha-50001 10 1,000,000 300 x 7.8 18342 £1,217

Alpha-60001 13 >10,000,000 300 x 7.8 18343 £1,217

Alpha-M (mixed bed)1 13 >10,000,000 300 x 7.8 18344 £1,217

SuperAW25002 4 5,000 150 x 6.0 19315 £1,219

SuperAW30002 4 90,000 150 x 6.0 19316 £1,219

SuperAW40002 6 1,000,000 150 x 6.0 19317 £1,219

SuperAW50003 7 1,000,000 approx. 150 x 6.0 19318 £1,219

SuperAW60003 9 10,000,000 approx. 150 x 6.0 19319 £1,219

SuperAWM-H3 9 10,000,000 approx. 150 x 6.0 19320 £1,2191 Use with Alpha guard column 18345 (£455). 13μm, 40 x 6.0mm 3 Use with SuperAW-H guard column 19322 (£399). 13μm, 35 x 4.6mm2 Use with SuperAW-L guard column 19321 (£399). 7μm, 35 x 4.6mm PEO = polyethylene oxide

Polymer-based Phases (TSKgel H Series)• Porous, cross-linked polystyrene-divinylbenzene• Chemically and thermally stable• Wide range of pore sizes, mixed bed columns and multipore phases

The TSKgel H type columns are based on porous, highly cross-linked, spherical polystyrene-divinylbenzene (PS-DVB) resin. These materials are available in 8 pore sizes and various particle sizes, and are compatible with non-polar organic solvents. They are suitable for the GPC analysis of oligomers, polymers and polymer additives. For polymer samples with a broad molecular weight range, mixed-bed columns are available.

TSKgel SuperMultiporeHZ SeriesThe SuperMultiporeHZ series comprises high performance organic SEC columns featuring a multipore gel with a wide distribution of pore diameters in each particle. Conventional SEC packing materials have a comparatively narrow pore size distribution, so for analyses of polymers of wide molecular weight distribution several columns may be connected in series or a single mixed-bed column used which contains several materials of different pore diameters (see Figure 4 on page 245). The TSKgel SuperMultiporeHZ columns produce calibration curves with excellent linearity (see Figure 6) and no inflection points, enabling more accurate determination of molecular weight.

TSKgel PhaseParticle

Size (μm)Exclusion Limit (polystyrene)

Analytical Column (150 x 4.6mm)

Guard Column (20 x 4.6mm)

£1,380 £478

SuperMultiporeHZ-H 6 40,000,000 21885 21886

SuperMultiporeHZ-M 4 2,000,000 21488 21489

SuperMultiporeHZ-N 3 1,200,000 21815 21816

Ordering Information – SuperMultiporeHZ SeriesColumn dimensions: 150 x 4.6mmEluent: THFFlow rate: 0.35ml/minDetection: UV, 254nmSample: Polystyrene

Figure 6. TSKgel SuperMultiporeHZ Series: Calibration curves

MW

Elution time (mins)




Hichrom Lim


• Silica- and polymer-based materials• Super series ideal for LC-MS• High recovery for biomolecules

Reversed-Phase Materials

Tosoh Bioscience offers a number of both silica-based and polymer-based columns for reversed-phase applications. The characteristics of these materials are summarised below.

TSKgel ODS-100V and ODS-100Z

• Ultra pure silica• TSKgel ODS-100V compatible with 100% aqueous eluent• Good retention of polar analytes

Silica-based Phases

TSKgel ODS-140HTPTSKgel® ODS-140HTP columns were developed for use in high throughput applications. They are packed with 2.3μm particles, providing high resolution and short analysis times at moderate pressure. The lower pressure drop enables these columns to be used with either UHPLC (up to 9,000psi) or conventional HPLC systems.

TSKgel ODS-100V and ODS-100Z reversed-phase columns are based on ultra pure silica with a high surface area (450m2/g). TSKgel ODS-100V provides strong retention for polar compounds due to its lower C18 ligand density (15% carbon content). The phase is prepared by an incomplete first reaction with a difunctional octadecylsilane reagent, followed by endcapping with a mixture of two difunctional dialkylsilane reagents. This enables the phase to provide complete wetting and retention stability in 100% aqueous eluents. Figure 7 shows the separation of 15 organic acids in less than 25 minutes using 0.1% phosphoric acid eluent.

TSKgel ODS-100Z contains a high density (20% carbon content) C18 bonded phase for maximum retention and selectivity of small molecular weight compounds. It is prepared by bonding the surface with a difunctional octadecylsilane, followed by repeated endcapping with monofunctional trimethylsilane reagent.

TSKgel PhaseFunctional

GroupBonding

Particle Size (μm)

Pore Size (Å)Carbon Load

(%)Endcapped

Exclusion Limit (Da)

Silica-based

ODS-140HTP C18 Polymeric 2.3 140 8 Yes 20,000

ODS-100V C18 Monomeric 3, 5 100 15 Yes 8,000

ODS-100Z C18 Monomeric 3, 5 100 20 Yes 8,000

ODS-80TM C18 Monomeric 5, 10 80 15 Yes 6,000

ODS-80TS C18 Monomeric 5, 10 80 15 Yes 6,000

Octyl-80TS C8 Monomeric 5 80 11 Yes 6,000

CN-80TS Cyano Monomeric 5 80 8 Yes 6,000

ODS-120T C18 Polymeric 5, 10 120 22 Yes 10,000

ODS-120A C18 Polymeric 5, 10 120 20 No 10,000

Super-ODS C18 Polymeric 2.3 110 8 Yes 10,000

Super-Octyl C8 Polymeric 2.3 110 5 Yes 10,000

Super-Phenyl Phenyl Polymeric 2.3 110 3 Yes 10,000

OligoDNA RP C18 Monomeric 5 250 10 No 165,000

TMS-250 C1 Monomeric 10 250 5 No 200,000

Protein C4-300 C4 Polymeric 3 300 3 Yes -

Polymer-based

Octadecyl-2PW C18 Monomeric 5 125 - - 8,000

Octadecyl-4PW C18 Monomeric 7, 13 500 - - 200,000

Phenyl-5PW RP Phenyl Monomeric 10, 13 1000 - - 1,000,000

Octadecyl-NPR C18 Monomeric 2.5 Non-porous - - >1,000,000

Column: TSKgel ODS-100V (5μm, 250 x 4.6mm)Eluent: 0.1% H

3PO

4, pH 2.3

Flow rate: 1.0ml/minTemperature: 40°C

Figure 7. Organic acids on TSKgel ODS-100V

1. Oxalic acid2. L-Tartaric acid3. Formic acid4. L-Malic acid5. L-Ascorbic acid6. Lactic acid7. Acetic acid8. Maleic acid

9. Citric acid10. Succinic acid11. Fumaric acid12. Acrylic acid13. Propionic acid14. Glutaric acid15. Itaconic acid

0 5 10 15 20 25Mins

1

24

3

5

6

7

8

109

11

12

13

14

15

TSKgel® Reversed-phase Materials

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Reversed-Phase Materials (continued)

TSKgel® Super SeriesThe silica particles used in the Super series columns are monodisperse spherical 2.3μm beads with 110Å pores. The Super-ODS, Super-Octyl and Super-Phenyl phases are polymerically bonded and exhaustively endcapped, to produce low metal-content, low silanol phases. The 2.3μm particle provides superior resolution and speed of analysis as well as improved sensitivity. The columns do not suffer from excessive pressure drops.

TSKgel® Super series columns provide high resolution separations for pharmaceuticals, peptides, nucleotides, amino acids and small proteins. In pharmaceutical stability testing, degradation products as low as 0.1% of the main component can be measured.

The high performance of TSKgel Super-ODS is shown by the rapid analysis of 18 PTC-derivatized amino acids in under 5 minutes (Figure 8).

Ordering Information – Silica-based Phases

TSKgel PhaseColumn Dimensions (mm)

50 x 2.1 100 x 2.1

ODS-140HTP 21927 £474 21928 £484

TSKgel Phase3μm

Column Dimensions1 (mm) Guard Cartridges2 (3/pk) (For 2.0mm i.d. columns)35 x 2.0 50 x 2.0 75 x 2.0 150 x 2.0

£311 £371 £381 £401 £227

100V 21813 21812 21811 21810 21997

100Z 22728 22729 22730 22732 21996

TSKgel Phase 5μm

Column Dimensions (mm) Guard Cartridges3 (3/pk) (For 4.6mm i.d. columns)50 x 2.0 150 x 2.0 150 x 4.6 250 x 4.6

£321 £344 £344 £388 £214

100V 21457 21458 21455 21456 21453

100Z 21460 21459 21461 21462 21454

TSKgel Phase5μm

Column Dimensions5 (mm)

50 x 2.0 100 x 2.0 150 x 2.0 250 x 2.0 50 x 4.6 75 x 4.6 100 x 4.6 150 x 4.6 250 x 4.6

- - £333 £388 - £309 - £333 £388

ODS-80TM - - - - - 16651 - 8148 8149

ODS-80TS - - 18150 18151 - 17200 - 17201 17202

Octyl-80TS - - - - - - - 17344 17345

CN-80TS - - - - - - - 17348 17349

- - £333 £388 - - - £383 £401

ODS-120T - - 18152 18153 - - - 7637 7125

ODS-120A - - - - - - - 7636 7124

£444 £456 - - £464 - £497 - -

Super-ODS4 19541 19542 - - 18154 - 18197 - -

£427 £439 - - £464 £509 £497 £509 -

Super-Octyl4 20013 20014 - - 18275 - 18276 - -

Super-Phenyl4 20017 20018 - - 18277 - 18278 - -

OligoDNA RP - - - - - - - 13352 -

TMS-2506 - - - - - 7190 - - -

- £428 £457 £505 - £428 - £457 £505 -

Protein C4-3007 22830 22831 22832 - 22827 - 22828 22829 -

1 Other dimensions available 2 Use with holder 19308 (£259) 3 Use with holder 19018 (£237). Other dimension guards available

4 2.3μm 5 Preparative dimensions and guard cartridges available 6 10μm particle size 7 3μm particle size

Column dimensions: 100 x 4.6mmEluent: CH

3CN - H

2O (60:40)

Flow rate: 1.5ml/minTemperature: Ambient

Figure 8. TSKgel Super-ODS for rapid separation of 18 PTC-derivatized amino acids

1 Asp 7. Thr 13. Met2. Glu 8. Ala 14. Cys3. Ser 9. Pro 15. Ile4. Gly 10.PTC-NH

2 16. Leu

5. His 11.Tyr 17. Phe6. Arg 12.Val 18. Lys

0 1 2 3 4 Mins

12

4

35

6

7

8

10

9 11

1213

141615 17

18

TSKgel Protein C4-300TSKgel Protein C4-300 is a 300Å pore size phase, designed for the optimal recovery and resolution of proteins such as recombinant proteins, antibody fragments or PEGylated proteins.




Hichrom Lim


Reversed-Phase Materials (continued)

Polymer-based PhasesThe polymer-based reversed-phase materials are chemically stable from pH 2 – 12, allowing operation at basic pH where silica-based phases have limited chemical stability. The non-porous Octadecyl-NPR column produces extremely fast kinetics and quantitative recovery of proteins at sub-microgram loads. Figure 9 shows the high resolution, fast separation of peptides on a TSKgel Octadecyl-NPR column.

TSKgel® Octadecyl-4PW is suitable for the analysis of peptides and small proteins, whereas the TSKgel Octadecyl-2PW is used for small pharmaceutical compounds at basic pH. TSKgel Phenyl-5PW has a high loading capacity and is ideal for the separation of high molecular weight proteins.

The specifications of these phases are summarised on page 247.

Ordering Information – Polymer-based Phases

TSKgel PhaseColumn Dimensions1 (mm) Guard Cartridge (3/pk)

75 x 2.0 150 x 2.0 35 x 4.6 75 x 4.6 150 x 4.6 150 x 6.0 For 2.0mm i.d. columns2

For 4.6mm i.d. columns3

Octadecyl-2PW - 18754 £538 - - 17500 £717 17501 £831 42161 £607 -

Octadecyl-4PW - 18755 £527 - - 13351 £499 - 42160 £298 19008 £214

Phenyl-5PW RP 18756 £469 - - 8043 £794 - - 42159 £336 19007 £214

Octadecyl-NPR - - 14005 £740 - - - - -1 Preparative dimensions available 2 Use with holder 19308 (£259) 3 Use with holder 19018 (£237)

• Ether, phenyl and butyl functionalities• Analytical and preparative columns• Wide pH range (2 – 12)• Fully scalable from analytical to preparative dimensions

The porous HIC packing materials are based on the G5000PW resin and derivatized with phenyl (Phenyl-5PW) or oligoethyleneglycol groups (Ether-5PW). The Ether-5PW is less hydrophobic than Phenyl-5PW and thus exhibits shorter retention times. Butyl-NPR is based on 2.5μm non-porous particles of the same chemical composition as G5000PW, and is useful for high speed applications. It is more hydrophobic than Phenyl-5PW. It also benefits from excellent mass recovery and is the preferred choice for process monitoring and quality control. The TSKgel HIC columns are compatible with water-soluble organic solvents at concentrations below 50% methanol, ethanol, acetonitrile, DMF, DMSO or below 30% chloroform.

Phase StructureParticle Size

(μm)Pore Size

(Å)

Ether-5PW 10, 13, 20 1000

Phenyl-5PW 10, 13, 20 1000

Butyl-NPR 2.5 -

Sample MW Range (Da) TSKgel Phase

Peptides <10,000 Butyl-NPR

Medium to large proteins

>10,000Phenyl-5PW, Ether-5PW,

Butyl-NPR

DNA, RNA and PCR products

>500,000 Phenyl-5PW, Butyl-NPR

Oligonucleotides >10,000 Phenyl-5PW, Butyl-NPR

TSKgel HIC Phases

Applications of TSKgel HIC Phases

Ordering Information – TSKgel HIC Phases

PhaseColumn Dimensions (mm) Guardgel Kit1

75 x 2.0 35 x 4.6 100 x 4.6 75 x 7.5 150 x 21.5 For 4.6 and 7.5mm i.d. columns

For 21.5mm i.d. columns

Ether-5PW (10μm) 18760 £824 - - 8641 £1,091 - 86433 £596 -

Phenyl-5PW (10μm) 18759 £824 - - 7573 £1,091 76562 £3,719 76523 £487 160952 £751

Butyl-NPR - 14947 £813 42168 £1,015 - - - -1 Kit contains guard, holder and connector 2 13μm material 3 20μm material

Hydrophobic Interaction (HIC) Phases

Column: TSKgel Octadecyl-NPR (35 x 4.6mm)Eluent: A: 100mM HClO

4

B: CH3CN - 100mM HClO

4 (80:20)

Gradient of A to B over 8 mins.Flow rate: 1.5ml/min

Figure 9. High speed analysis of peptides

1. Met-enkephalin2. Leu-enkephalin3. Oxytocin4. Bombesin5. Somatostatin6. Calcitonin7. b-Endorphin

0 2 4 6 Mins

1 2

4

3

5

6

7


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• Covalently bonded carbamoyl and amino phases• Compatible with LC-MS• TSKgel® Amide-80 shows unique retention mechanism for saccharide analysis

Hydrophilic Interaction (HILIC) Phases

TSKgel® Amide-80 columns are well documented in the literature for HILIC separations of polar compounds. The phase consists of spherical silica particles that are covalently bonded with non-ionic carbamoyl groups. Columns can be operated over a wide range of eluent conditions for use with a broad range of sample polarities. Target applications include the analysis of hydrophilic biomolecules, including saccharides, glycans, oligosaccharides, peptides, nucleic acids and small molecules for drug discovery, that are weakly retained by reversed-phase chromatography. Figure 10 shows the separation of b-cyclodextrin hydrolysate on a 5μm TSKgel Amide-80 column.

The TSKgel NH2-100 phase is based on silica treated with a special endcapping procedure followed by the stepwise introduction of amino groups. Due to the high ligand density and large surface area, 3μm TSKgel NH2-100 columns show high retention for very polar compounds, including carbohydrates, peptides, vitamins and polar drugs. In addition, they have been shown to exhibit increased long term stability compared with conventional amino phases.

The selectivity of TSKgel NH2-100 differs from TSKgel Amide-80 as shown in Figure 11. If selectivity or regulatory requirements do not limit column choice, then TSKgel Amide-80 is generally preferred over amino phases due to greater long term stability.

Structure of TSKgel Amide-80

Ordering Information – HILIC Phases

Phase 3μmColumn Dimensions (mm) Guard Cartridges (3/pk)

50 x 2.0 150 x 2.0 50 x 4.6 150 x 4.6 For 2.0mm i.d. Columns1 For 4.6mm i.d. Columns2

Amide-80 21864 £585 21865 £633 21866 £580 21867 £648 21862 £235 21863 £212

NH2-100 21967 £464 21968 £516 21969 £464 21970 £516 21971 £206 21972 £206

Phase 5μmColumn Dimensions3 (mm) Guard Cartridges1

(3/pk)50 x 2.0 100 x 2.0 150 x 2.0 250 x 2.0

Amide-80 19694 £569 19695 £609 19696 £615 19697 £627 21941 £237

Phase 5μmColumn Dimensions3 (mm)

Guard ColumnGuard Cartridges2

(3/pk)50 x 4.6 100 x 4.6 250 x 4.6

Amide-80 19532 £563 19533 £608 13071 £666 19021 £186 19010 £2141 Use with holder 19308 (£259) 2 Use with holder 19018 (£237) 3 Other dimensions, including preparative, available

Sample: ß-cyclodextrin hydrolysate, 1-7 degrees of polymerizationColumn: 250 x 4.6mm (5μm)Eluent: CH

3CN - H

20 (55:45)

Flow rate: 1.0ml/minDetection: RI

Figure 10. Separation of ß-cyclodextrin hydrolysate on TSKgel Amide-80

Figure 11. Separation of PA-glycans

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Column: (a) TSKgel NH2-100 (3μm, 150 x 4.6mm) (b) TSKgel Amide-80 (3μm, 150 x 4.6mm) Eluent: (a) A: 0.2M TEA acetate, pH 6.5 – CH

3CN (30:70)

B: 0.2M TEA acetate, pH 6.5 – CH3CN (60:40)

(b) A: 0.2M TEA acetate, pH 6.5 – CH3CN (26:74)

B: 0.2M TEA acetate, pH 6.5 – CH3CN (50:50)

Flow rate: 1.0ml/minGradient: 0 – 100% in 30 mins (hold 15mins)Detection: Fluorescence Ex 315nm, Em 380nmTemperature: 40°C

10 15 20 25 30 35 Mins

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TSKgel Phase Functional Group Particle Size (μm) Pore Size (Å)Surface Area

(m2/g)Temperature Range (°C)

pH Range

Amide-80 Carbamoyl 3, 5 100 4504 - 80 (5μm) 4 - 40 (3μm)

2.0 - 7.5

NH2-100 Ethylamino 3 100 450 10 - 50 2.0 - 7.5




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• Silica and polymer based anion and cation exchange materials• Range of pore sizes• Porous and non-porous materials

Ion-Exchange Phases

Tosoh Bioscience offers a range of silica and methacrylate based high efficiency columns for analysis and isolation of biomolecules by anion and cation exchange chromatography. The BioAssist materials were created to provide higher capacity for larger proteins and have biocompatible PEEK hardware.

TSKgel® Ion-Exchange Phases

TSKgel Phase Type Functional Group pH Range Particle Size (μm) Pore Size (Å)Exclusion Limit

(Da) for PEG

Silica-based (SW)

DEAE-2SW WAX DEAE 2 - 7.5 5 125 10,000

DEAE-3SW WAX DEAE 2 - 7.5 10 250 30,000

CM-2SW WCX Carboxymethyl 2 - 7.5 5 125 10,000

CM-3SW WCX Carboxymethyl 2 - 7.5 10 250 30,000

SP-2SW SCX Sulphopropyl 2 - 7.5 5 125 10,000

Polymer-based (PW)

SuperQ-5PW SAX Trimethylamino 2 - 12 10, 13 1000 1,000,000

DEAE-5PW WAX DEAE 2 - 12 10, 13, 20 1000 1,000,000

BioAssist Q SAX Polyamine 2 - 12 10, 13 ∼4000 >5,000,000

SP-5PW SCX Sulphopropyl 2 - 12 10, 13, 20 1000 1,000,000

CM-5PW WCX Carboxymethyl 2 - 12 10, 13 1000 1,000,000

BioAssist S SCX Sulphopropyl 2 - 12 7, 13 ∼1300 ∼4,000,000

Non-porous (NPR)

DEAE-NPR WAX DEAE 2 - 12 2.5 ∼0 500

SP-NPR SCX Sulphopropyl 2 - 12 2.5 ∼0 500

Q-STAT SAX Trimethylamino 3 - 10 7, 10 ∼0 500

DNA-STAT SAX Trimethylamino 3 - 10 5 ∼0 500

CM-STAT WCX Carboxymethyl 3 - 10 7, 10 ∼0 500

SP-STAT SCX Sulphopropyl 3 - 10 7, 10 ∼0 500

Silica-based Phases (SW)Silica-based ion-exchange materials have a smaller pore size compared to the polymer-based phases, and are most suitable for analysing smaller molecular weight samples such as nucleotides, pharmaceuticals, catecholamines and small peptides.

Polymer-based Phases (PW)The porous methacrylate based phases are derived from G5000PW (10μm, 1000Å) spherical particles, derivatised with DEAE, SP or CM functionalities. The SuperQ-5PW is a higher capacity strong anion- exchanger, as a result of the polyamine functional groups. All PW phases show excellent stability from pH 2–12 and offer good mass and activity recovery for enzymes, proteins, DNA, nucleic acids and polypeptides.

Each PW phase has an equivalent in 20 and 30μm particle size TSKgel 5PW resins and even larger particle TOYOPEARL process media. This means that a method developed on an analytical column can be reliably scaled up to preparative and process scale dimensions, using the same backbone chemistry.

Particles in the TSKgel BioAssist Q columns contain very large pores (∼4000Å) that are functionalised with polyamine groups to form a network structure. The dynamic binding capacity of BioAssist Q columns is high over a wide molecular weight range. TSKgel BioAssist S columns are packed with polymethacrylate particles possessing 1300Å pores functionalised with sulphopropyl groups. Columns are supplied in biocompatible PEEK hardware for analysis by HPLC or FPLC. TSKgel BioAssist phases are particularly suitable for the analysis of monoclonal antibodies, plasmids and other large proteins.

Column: TSKgel DEAE-5PW (75 x 7.5mm)Eluent: A. 0.1M Tris-HCl buffer, pH 7.6 + 0.3M NaCl B. A + 0.1M NaClGradient: A to B linear gradient over 300 minsFlow rate: 1ml/min

Figure 12. Separation of E. coli RNA

4S tRNA

5S rRNA

23S rRNA

16S rRNA

High MW Impurities

0 40 80 Mins

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Ion-Exchange Phases (continued)

Non-Porous Resin (NPR) MaterialsThe NPR ion-exchange materials such as DEAE-NPR, DNA-NPR and SP-NPR are based on methacrylate polymer of 2.5μm particle size. High efficiency, coupled with low sample capacity, makes these columns suitable for fast analysis and micro-scale preparative isolation of DNA digests, and haemoglobin variants.

TSKgel STAT SeriesTSKgel STAT columns, the newest series of non-porous ion-exchange columns, enable high performance separations with short analysis times. These phases consist of monodisperse, non-porous polymethacrylate resin particles, the surface of which consists of an open access network of multi-layered anion- or cation-exchange groups. The innovative bonding chemistry, combined with a relatively large particle size, results in a high loading capacity and a lowoperating pressure, compared with traditional non-porous resins.

Table 1 illustrates that despite the fact that surface area decreases with increasing particle size, the larger TSKgel Q-STAT and DNA-STAT particles have higher binding capacities than the smaller particles used in TSKgel NPR columns. The novel bonding chemistry results in a significant increase in static binding capacity, more than compensating for the loss in external surface area of the larger particles.

Schematic diagram of TSKgel STAT series

Table 1

PropertyTSKgel

DEAE-NPRTSKgel

DNA-STATTSKgel Q-STAT

Particle Size 2.5μm 5μm 7μm 10μm

Capacity* 9.1 38.6 27.0 20.9

* Static binding capacity in mg BSA/mg dry gel

TSKgel STAT 10μm particle size columns in 35 x 3.0mm column dimensions are ideally suited for rapid candidate screening or process monitoring. The smaller 7μm particle size columns (100 x 4.6mm) are designed for high resolution separations of nucleic acids, mAb variants, PEGylated protein or protein aggregates.

Figure 13 shows the reaction monitoring for the PEGylation of lysozyme at 4 minute intervals on a TSKgel SP-STAT column. As demonstrated in the chromatogram, peak areas of mono-, di- and tri-PEGylated lysozyme increased with reaction time, whilst the area of unreacted lysozyme decreased. Figure 14 illustrates the utility of TSKgel DNA-STAT for the separation of mono-, di- and tri-nucleotides.

0 0.5 1.0 Mins

Figure 13. Monitoring PEGylation reaction of lysozyme

Column: TSKgel DNA-STAT (5μm, 100 x 4.6mm)Eluent: A: 20mM Tris-HCl, pH 8.5 B: 0.75mM NaCl in buffer AGradient: 50 to 75% B in 25 minsFlow rate: 0.8ml/minDetection: UV, 260nm

Column: TSKgel SP-STAT (10μm, 35 x 3.0mm)Eluent: A: 20mM acetate buffer, pH 5.0 B: 0.5M NaCl in A, pH 5.0Gradient: 0 to 100% B in 1.5 minsFlow rate: 2.0ml/minDetection: UV, 280nm

Figure 14. Separation of nucleotides on TSKgel DNA-STAT

0 5 10 15 20 25Mins

Native

Mono-PEG

Di-PEG

Tri-PEG



Hichrom Lim


Ion-Exchange Phases (continued)

Ordering Information – Ion-Exchange Phases

TSKgel Phase Column Dimensions (mm) Guardgel Kit1

(For 4.6 and 7.5mm i.d. Columns)250 x 2.0 250 x 4.6 75 x 7.5

DEAE-2SW 18761 £807 7168 £970 - 76482 £507

DEAE-3SW - - 7163 £970 76482 £507

CM-2SW - 7167 £970 - 76502 £507

CM-3SW - - 7162 £970 76502 £507

SP-2SW - 7165 £1,131 - -

TSKgel PhaseColumn Dimensions (mm) Guardgel Kit1

75 x 2.0 35 x 4.6 50 x 4.6 75 x 7.5 150 x 21.5 For 4.6 and 7.5mm i.d. Columns

For 21.5mm i.d. Columns

SuperQ-5PW - - - 182573 £1,315 183875 £5,215 18388 £562 183902 £1,269

DEAE-5PW 187573 £807 - - 71643 £1,073 75745 £3,748 7210 £562 160922 £751

BioAssist Q - - 196854 £1,021 - - - -

SP-5PW 187583 £807 - - 71613 £1,073 75755 £3,748 7211 £562 160932 £751

CM-5PW - - - 130683 £1,073 - 13069 £536 -

BioAssist S - - 196864 £1,021 - - - -

DEAE-NPR - 13075 £813 - - - - -

SP-NPR - 13076 £813 - - - - -1 Kit contains guard, holder and connector 2 20μm material 3 10μm material 4 PEEK hardware 5 13μm material

TSKgel Phase Particle Size (μm) Column Dimensions (mm) Cat. Number Price

Q-STAT10 35 x 3.0 21960 £722

7 100 x 4.6 21961 £1,082

DNA-STAT 5 100 x 4.6 21962 £1,082

CM-STAT10 35 x 3.0 21965 £722

7 100 x 4.6 21966 £1,082

SP-STAT10 35 x 3.0 21963 £722

7 100 x 4.6 21964 £1,082

Please contact Hichrom to request a copy of the Tosoh Bioscience Chromatography catalogue or any product brochure.


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• Choice of 3 affinity ligands• Rigid polymeric base resin• Products for analytical, preparative through to process scale• Wide pH range

Affinity Chromatography Phases

Affinity chromatography offers the highest level of specificity and selectivity in biomolecular separations and purifications. TSKgel® affinity columns are based on G5000PW (5PW) porous resin and are available in two group-specific stationary phases and one activated packing (Tresyl-5PW). Process scale media, based on TOYOPEARL HW-65 resin (65μm) made by the same polymer chemistry, enable easy scale-up. The choice of a specific ligand is determined by the expected interaction between sample and column bonded phase.

TSKgel® Affinity PhasesTSKgel Affinity Phase Ligand Type Particle Size (μm) Pore Size (Å) pH Range Adsorption Capacity

Boronate-5PWm-Aminophenyl

boronate10 1000 2 - 9

40μmol/ml resin for sorbitol

Chelate-5PW Iminodiacetic acid 10 1000 2 - 12 -

Tresyl-5PW Tresyl 10 1000 2 - 12>60mg/g dry resin for

soybean trypsin inhibitor

Boronate-5PWUnder alkaline conditions the boronate anionic structure of the TSKgel Boronate-5PW can bind with 1,2 cis-diol groups, such as those in carbohydrates, carbohydrate-containing compounds and catecholamines. This interaction is enhanced in the presence of Mg2+ ions and inhibited by amine-containing buffers.

Chelate-5PWTSKgel Chelate-5PW utilizes the ability of iminodiacetic acid to chelate ions such as Zn2+ and Cu2+. Peptides and proteins containing histidine residues can adsorb to these chelated ions at neutral pH and then be eluted with buffer. Applications for Chelate-5PW include immunoglobulins, transferrins, lectins, milk proteins and membrane proteins. Figure 15 shows the separation of porcine pancreatic elastase on a TSKgel Chelate-5PW column loaded with copper ions.

Tresyl-5PWThe Tresyl-5PW (2,2,2-trifluoroethanesulphonyl) requires activation with a user-selected ligand containing amino, thiol, phenol or imidazole groups. This results in a ‘custom’ affinity ligand. Principal applications include the selective purification of antigens after coupling of the appropriate antibody.

TSKgel PhaseColumn Dimensions (mm) Guardgel Kit1

(For 7.5mm i.d. Columns)40 x 6.0 75 x 7.5

Boronate-5PW - 13066 £1,014 13125 £615

Chelate-5PW - 8645 £1,028 8647 £615

Tresyl-5PW 14455 £1,180 14456 £1,352 -

Ordering Information – Affinity Phases

1 20μm material. Kit contains Guardgel, holder and connector

TSKgel Chelate-5PW TSKgel Tresyl-5PWTSKgel Boronate-5PW

Column: TSKgel Chelate-5PW (75 x 7.5mm) Column loaded with copper ionsEluent: A: 0.5M NaCl in 20mM phosphate buffer, pH 7.0 B: A + 20mM imidazole Linear gradient A to B over 30 minsFlow rate: 1ml/min

Figure 15. Separation of porcine pancreatic elastase by immobilized metal ion affinity chromatography

0 10 20 30Mins




Hichrom Lim


• TOYOPEARL and TSKgel® resins• Seamless scale-up from analytical to process scale• Ideal for peptide, protein and oligonucleotide separations• Prepacked columns for scouting and method development

Bulk Resins

Tosoh Bioscience offers a comprehensive range of media for all common modes of liquid chromatography, including ion-exchange, hydrophobic interaction, reversed-phase, size exclusion and affinity.

TOYOPEARL resins are hydrophilic, macroporous methacrylate-based media for large-scale medium pressure liquid chromatographic applications. They exhibit high mechanical and chemical stability and are stable from pH 2 – 12 for normal operating conditions and pH 1 – 13 for cleaning conditions. The resins are available in average particle sizes of 35, 65, 75 and 100μm.

TSKgel® PW resins are larger particle size versions (20 and 30μm) of the methacrylate packing materials used for ion-exchange and hydrophobic interaction chromatography columns. The high degree of cross-linking leads to high efficiency purifications.

Method DevelopmentLABPAK kits containing small volumes of bulk resins of various functionalities are available for method development. In addition, ToyoScreen prepacked columns for a range of TOYOPEARL products are recommended (see page 257).

IEC Bulk Resins• Weak and strong anion and cation exchangers• Temperature stable from 4 - 60°C (autoclavable at 120°C)• Suitable for pH range 2 - 13

SEC Bulk Resins• 5 pore sizes and 3 particle sizes (S, F, C)• HW-40 ideal for desalting applications

HIC Bulk Resins• Wide range of hydrophobicities• Standard 1000Å pore size

Ion Exchange Resin TypeAverage

Particle Size (μm)Pore Size

(Å)

TSKgel

DEAE-5PW WAX 20, 30 1000

SuperQ-5PW SAX 20, 30 1000

SP-5PW SCX 20, 30 1000

SP-3PW SCX 30 250

TOYOPEARL1

DEAE-650 WAX 35, 65, 100 1000

SuperQ-650 SAX 35, 65, 100 1000

QAE-550 SAX 100 500

GigaCap Q-650 SAX 75 1000

Q-600C AR SAX 100 750

CM-650 WCX 100 1000

SP-550 SCX 100 500

SP-650 SCX 35, 65, 100 1000

MegaCap ll SP-550 SCX 200 500

GigaCap S-650 SCX 75 1000

GigaCap CM-650 WCX 75 10001 35μm = S grade, 65μm = M grade, 100μm = C grade, 200μm = EC grade

1 35μm = S grade, 45μm = F grade, 75μm = C grade 2 Determined using globular proteins

1 35μm = S grade, 65μm = M grade, 100μm = C grade2 mg lysozyme/ml resin

TOYOPEARL Resin1

Average Particle Size (μm)

Pore Size (Å) Exclusion Limit (Da)2

HW-40 35, 45, 75 50 1 x 104

HW-50 35, 45 125 8 x 104

HW-55 35, 45 500 7 x 105

HW-65 35, 45 1000 5 x 106

HW-75 45 >1000 5 x 107

HIC Resin1 Average Particle Size (μm)

Adsorption Capacity2

TSKgel Ether-5PW 20, 30 10-30

TSKgel Phenyl-5PW 20, 30 10-30

TOYOPEARL Ether-650 35, 65 10-30

TOYOPEARL Phenyl-650 35, 65, 100 30-50

TOYOPEARL Butyl-650 35, 65, 100 30-50

TOYOPEARL Hexyl-650 100 30-50

pKa values for Ion Exchange groups

Q, SuperQ, QAE pKa = 12.2

CM pKa = 4.7*

*pKa Toyopearl GigaCap CM-650M = 3.6

S, SP pKa = 1.2

DEAE pKa = 11.5

Cation

Anion7.0

Decreasing resin pKa

pKa values for Ion Exchange groups

CM pKa =4.7*

S, SP pKa =1.2

Q, SuperQ, QAE pKa =12.2

DEAE pKa =11.5

*pKa TOYOPEARL GigaCap CM-650M = 3.6

Please contact Hichrom for ordering information on Tosoh Bioscience bulk resins

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Affinity Chromatography Bulk Resins• Active, reactive and group specific resins• Standard 1000Å pore size• Efficient antibody purification• New high capacity AF-rProtein A resin

Bulk Resins (continued)

TOYOPEARL AF-rProtein A-650FTOYOPEARL AF-rProtein A-650F is an affinity resin designed for efficient and robust large-scale purification of monoclonal antibodies (mAb). A newly developed recombinant Protein A ligand, derived from one of the IgG-binding domains of the staphylococcus aureus Protein A, is linked to the well proven methacrylic polymer backbone.

Multipoint attachment of the ligand to the TOYOPEARL matrix further enhances the chemical and thermal stability of the resin. In practice this leads to a low level of Protein A leaching and also to a high resistance to alkaline solutions employed in cleaning-in-place (CIP) procedures.

TOYOPEARL AF-rProtein A-650F binds human and mouse immunoglobulin G, as well as immunoglobulin M and Fab fragments. Key features of this product include:

• Efficient mAb capturing from high titre feedstocks• High IgG binding capacity over a wide range of velocities• High alkaline stability for effective cleaning and sanitization• Rigid polymethacrylate base matrix with ideal pressure/flow characteristics

Figure 16 shows the purification of a humanized IgG1 from a Chinese Hamster Ovary (CHO) cell lysate. Remaining host cell proteins and leached Protein A ligand amounts were very low, resulting in very high product purity.

Please contact Hichrom for ordering information on Tosoh Bioscience bulk resins.

TOYOPEARL Resin Resin Type Average Particle Size (μm)

AF-rProtein A-650F Protein A Affinity 45

AF-Tresyl Activated resin 65

AF-Epoxy Activated resin 65

AF-Amino Reactive resin 65

AF-Carboxy Reactive resin 65

AF-Formyl Reactive resin 65

AF-Blue HC Group specific 65

AF-Red Group specific 65

AF-Chelate Group specific 65

AF-Heparin HC Group specific 65

Recombinant Protein A Derived Ligand Figure 16. Purification of humanized IgG1

Column: TOYOPEARL AF-rProtein A-650F (49 x 5mm)Binding buffer: 20mM Na phosphate, 150mM NaCl, pH 7.4Elution buffer: 100mM glycine-HCl, pH 3.0Flow rate: 0.5ml/minSample: 40ml CHO cell culture lysate containing 0.5mg/ml humanized IgG1

0 10 20 30 40 50Elution volume (mL)

600

50

100

B%



Hichrom Lim


• Prepacked cartridge design columns• 1ml and 5ml columns• Easy connections with AKTA®, FPLC and HPLC systems• Short scouting runs• Excellent recovery• Range of TOYOPEARL resins

ToyoScreen® Process Development Columns

ToyoScreen® columns for early process development are available in a range of TOYOPEARL resins including hydrophobic interaction, ion-exchange and affinity resins, with both 1ml and 5ml resin volumes available. They are simple, cartridge-type pre-packed columns, designed for small scale purification and as a convenient means of performing early resin screening for both target retention and recovery. Multiple columns can be connected in series for additional separation. They are easy to use and can be connected to most laboratory chromatography systems, requiring only a reusable ToyoScreen column holder.

All ToyoScreen columns are packed with TOYOPEARL materials which are also readily available as bulk polymeric media for scale-up and production. Figure 17 shows the different selectivities obtained for a standard protein mixture using the six different ToyoScreen (1ml) HIC columns.

Ordering Information (6/pk)

ToyoScreen HIC PhasesToyoScreen HIC Phase

1ml1 5ml1

£132 £289

Ether-650M 21372 21373

Phenyl-650M 21374 21375

Butyl-650M 21376 21377

Hexyl-650C 21378 21379

PPG-600M 21380 21381

Phenyl-600M 21892 21893

Butyl-600M 21494 21495

SuperButyl-550C 21382 21383

HIC Mix Pack2 21398 21399

ToyoScreen Affinity PhasesToyoScreen Affinity Phase

1ml1 5ml1

£145 £427

AF-Chelate-650M 21384 21385

AF-Blue HC-650M 21386 21387

AF-Red-650M 21388 21389

AF-Heparin HC-650M 21390 21391

£690 £2,070

AF-rProtein A-650F 228093 228113

ToyoScreen Ion-exchange PhasesToyoScreen Ion-exchange Phase

1ml1 5ml1

£95 £201

DEAE-650M 21360 21361

SuperQ-650M 21362 21363

QAE-550C 21364 21365

Q-600C AR 21992 21993

GigaCap Q-650M 21859 21860

CM-650M 21366 21367

SP-650M 21368 21369

SP-550C 21370 21371

MegaCap ll SP-550EC 21870 21871

GigaCap S-650M 21868 21869

IEC Anion Mix Pack4 21392 21393

IEC Cation Mix Pack5 21394 21395

IEC Mix Pack6 21396 21397

1 Use with ToyoScreen column holder 21400 (£37)2 1 each of PPG-600M, Butyl-600M, Phenyl-650M, Butyl-650M, Phenyl-600M, Hexyl-650C3 5/pk4 1 each of DEAE-650M, SuperQ-650M, QAE-550C, GigaCap Q-650M, Q-600C AR5 1 each of CM-650M, SP-650M, SP-550C, GigaCap CM-650M, GigaCap S-650M6 1 each of GigaCap Q-650M, GigaCap S-650M, GigaCap CM-650M, Q-600C AR, SuperQ-650M, SP-550C

Figure 17. Screening of TOYOPEARL HIC resins – standard proteins

Column: ToyoScreen (1ml)Eluent A: 0.1M Phosphate buffer + 1.8M Na

2SO

4 (pH 7)

Eluent B: 0.1M Phosphate buffer (pH 7)Flow rate: 1ml/minGradient: 30 min linearInjection volume: 50µlSamples: Ribonuclease A, Lysozyme, α-Chymotrypsinogen (1mg/ml)

Ether-650M

PPG-600M

Phenyl-650M

Butyl-650M

SuperButyl-550C

Hexyl-650C

0 10 20 30 40 50 Mins


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