IEX Selection Kit 5 mL Column Product Code: 6609 Includes: 6601, 6603, 6605 & 6607
INTRODUCTION Properties of the 5 mL Ion Exchange columns: 5 mL axial flow column (ID: 1.6 cm diameter x 2.5 cm bed height)
COLUMN: COLUMN MATERIAL:
Polypropylene
LIGAND:
Sulphonic acid with a propyl linker (SP)
Carboxymethyl (CM)
Quaternary ammonium (Q)
Diethylamino ethyl (DEAE)
TOTAL IONIC CAPACITY:
≥ 175 μmol/g settled gel
90 to 130 μmol/g settled gel
40 - 60 μmol/g settled gel
110 to 160 μmol/g settled gel
BINDING CAPACITY:
≥ 90 mg/mL of adsorbent (lysozyme)
≥ 60 mg/mL of adsorbent (lysozyme)
75 - 100 mg/g of adsorbent (BSA)
76 - 120 mg/g of adsorbent (BSA)
MEAN PARTICLE SIZE (µm)
90 ± 10 Cross-linked 6% near-monodisperse agarose (PuraBead® 6HF)
MATRIX: MAXIMUM RECOMMENDED OPERATING PRESSURE (DELTA COLUMN PRESSURE):
Not exceeding 5 bar (~72 psi)*
RECOMMENDED OPERATIONAL FLOW RATE:
Up to 5 mL/min
OPERATING PH:
pH 4.0 to pH 14.0
PH STABILITY:
Long term: pH 3.0 – pH 12.0
CHEMICAL STABILITY:
All commonly used aqueous buffers and solutions
CLEANING/SANITIZATION:
STORAGE:
pH 2.0 to pH 14.0
0.5 to 1.0 M NaOH, 25 °C 2 – 8°C**, 20% ethanol, 0.2 M sodium acetate
* Delta pressure across the column (not the system pressure) ** To minimize the risk of the columns drying out.
IEX Selection Kit 5 mL Column PC6609 User Guide-v3
2 – 8°C**, 20% ethanol
INTRODUCTION Anion exchange adsorbents have been designed for capture, intermediate or polishing steps for the purification of negatively charged biom olecules, whereas Cation exchange adsorbents have been designed for capture and intermediate steps for the purification of positively charged biomolecules. Q PuraBead® HF is a high-performance strong anion exchange chromatography adsorbent with a quaternary ammonium group (Q). DEAE PuraBead® HF is a high-performance weak anion exchange chromatography adsorbent with a diethylamino ethyl group. SP PuraBead® HF is a high-performance strong cation exchange chromatography adsorbent with a Sulphonic acid group coupled via a propyl linker (SP). CM PuraBead® HF is a high-performance weak cation exchange chromatography adsorbent with a carboxy methyl group. All ligands are coupled to highly cross-linked near monodisperse 6% beaded agarose (PuraBead® 6HF) which has excellent flow properties. All IEX adsorbents are stable in up to 1.0M sodium hydroxide which allows for stringent cleaning and sanitization protocols. The pre-packed IEX columns are constructed using polypropylene and have been specifically designed to be cost effective, robust and provide a platform for fast and accurate process development and polishing steps for the binding and eluting of proteins using ion-exchange chromatography.
IEX Selection Kit 5 mL Column PC6609 User Guide-v3
OPERATING INSTRUCTIONS 1. The preferred option is to use the IEX 5 mL column with a liquid chromatography system or automated workstation. Note: The column can also be operated manually using a peristaltic pump or even a syringe. 2. Recommended equilibration buffers: • For CIEX: 20 mM phosphate pH 6.0. Sodium citrate and sodium acetate buffer can also be used. • For AIEX: 25 mM Tris buffer pH7.5. Other buffers suitable for use include Sodium phosphate, sodium citrate and HEPES. Note: IEX adsorbents are designed for adsorption of positively (pH < pI) or negatively charged proteins (pH > pI). It is recommended to use an equilibration buffer with a pH of at least 0.5 units below the iso-electric point (pI) of the target protein for CIEX and an equilibration buffer with a pH of at least 0.5 units above the iso-electric point (pI) of the target protein for AIEX. The ionic strength of the equilibration buffer should also be low, with preferably no or minimal salt present. 3. Allow the column, buffers and sample to reach the operational temperature. 4. Remove the inlet (Top) stopper of the column and attach directly to the liquid chromatography system using a standard 10-32 threaded male for 1/16’’ OD tubing connector, ensuring that the tubing connecting the workstation to the column is primed with equilibration buffer. 5. Snap off the outlet (Bottom) plug of the column and attach to the workstation as in step 4. Note: The reverse of the bottom plug is threaded and can be used as a stopper.
Column Inlet (TOP)
Column Outlet (BOTTOM)
6. Apply the protein solution/sample onto the column at a flow rate of 0.5 mL/min (2-minute residence time). 7. Remove any non-bound material in the column with up to 5 CV of equilibration buffer, or until the UV trace returns to baseline. 8. Elute the bound protein by increasing the conductivity of the solution with up to 5 CV of elution buffer. Any of the recommended equilibration buffers (for the respective IEX with the addition of up to 1.0 M sodium chloride (NaCl) would be suitable. For initial investigations, it is recommended to carry out a salt elution gradient (e.g. 20 CV from 0 to 1.0 M NaCl in equilibration buffer) to determine the appropriate elution
IEX Selection Kit 5 mL Column PC6609 User Guide-v3
condition for your target biomolecule and will also identify a purification strategy (i.e. separation of non-target proteins). 9. If a CIP is required, use up to 5 CV of 0.5 to 1.0 M NaOH. A contact time of 1 hour will normally suffice to ensure destruction of viable organisms, although up to 5 hours contact time may be required. No less than 5 column volumes are recommended. 10.Re-equilibrate column with up to 5 CV of equilibration buffer (to remove sodium hydroxide) and check pH and conductivity of the column eluate is equal to that of the buffer entering the column before storage or re-use. 11.If the column is to be stored for future use, place the column into the storage solution and store at 2 – 8 °C (to minimize the risk of the columns drying out). Note: The columns cannot be opened or re-packed.
IEX Selection Kit 5 mL Column PC6609 User Guide-v3
ORDER INFORMATION Gel Slurry Code
Description
Pack Size
6600
Q PuraBead® HF
4 x 1 mL
6601
Q PuraBead® HF
4 x 5 mL
6602
DEAE PuraBead® HF
4 x 1 mL
6603
DEAE PuraBead® HF
4 x 5 mL
6604
SP PuraBead® HF
4 x 1 mL
6605
SP PuraBead® HF
4 x 5 mL
6606
CM PuraBead® HF
4 x 1 mL
6607
CM PuraBead® HF
4 x 5 mL
6608
IEX Selection Kit (1mL columns)
1x 1mL Q PuraBead® HF 1x 1mL DEAE PuraBead® HF 1x 1mL SP PuraBead® HF 1x 1mL CM PuraBead® HF
6609
IEX Selection Kit (5mL columns)
1x 5mL Q PuraBead® HF 1x 5mL DEAE PuraBead® HF 1x 5mL SP PuraBead® HF 1x 5mL CM PuraBead® HF
We also offer a range of larger pack sizes for supply of bulk resins into cGMP development and manufacturing scale processes. For more information on this or any other supply related matters please do not hesitate to contact us on sales@astrea-bio.com
IEX Selection Kit 5 mL Column PC6609 User Guide-v3
Issue Date: 31 May 2022 CCR Number: CCR-1729 Author Name: R Dodd QA Reviewer Name: R Hawkins This product is covered by or for use under one or more patents: www.astreabioseparations.com/patents All trademarks, trade names, trade dress, product names and logos appearing on this document are the property of Astrea UK Services Ltd. Copyright © 2022 Astrea Bioseparations Ltd. All rights reserved.