Labeled protein purification resin solutions
1. Background
Ni-IDA Pharose FF is a metal chelating resin containing iminodiacetic acid (IDA) groups, which uses agarose microspheres as a matrix to activate the coupling of iminodiacetic acid (IDA) and then chelate Ni2+. It is suitable for affinity chromatography of most histidine-tagged (His-tag) proteins.
Ni-NTA Sepharose FF is a metal chelating filler containing a nitritriacetic acid (NTA) group, which uses agarose microspheres as a matrix to activate the coupling of nitritriacetic acid (NTA) and then chelate Ni2+. Compared with iminodiacetic acid (IDA), it has better tolerance to chelating agents, reducing agents, alkaline. It is mainly used for affinity chromatography of histidine-tagged (His-tag) proteins.
Figure 1 Flow chart of the use of Ni-IDA agarose gel FF series
Fig.2 Flow chart of the use of Ni-NTA agarose gel FF series
2. Parameters of tagged protein purification resins
|
category |
Ni-IDA agarose gel FF |
Ni-NTA agarose gel FF |
|
matrix |
6% cross-linked agarose |
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|
Ligands |
Iminodiacetic acid, ~30μmol Ni2+/mL |
Hypotritriacetic acid, ~17μmol Ni2+/mL |
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Particle size rangea |
45~165μm |
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Average particle size |
~90μm |
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Dynamic binding capacityb |
≥ 40 mg/mL histidine-tagged protein |
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Recommended workflow speed |
60~300cm/h |
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Maximum flow rate and pressure |
＞1500cm/h，0.5MPa |
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Use pH |
4~8.5 (recommended working pH), 3~12 (long-term stability); 2~14(Short-term stability) |
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Chemical stability |
It is stable in the following solutions: commonly used aqueous buffer, 1mol/L sodium hydroxide, 8mol/L urea, 6mol/L guanidine hydrochloride, 70% ethanol, etc |
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Concentrate:
a: Microspheres with more than 90% volume in this particle size range;
b: DBC10% test conditions were: 10% flow, E. coli expression of histidine-tagged recombinant protein, 6min residence time.
3. Application data:
Figure 3 Lane M: Marker
Lane 1 ：Start material
Lane 2: Flowthrough 1
Lane 3: Flowthrough 2
Lane 4: Elution (20 mM Imidazole)
Lane 5: Elution (50 mM Imidazole)
Lane 6: Elution (100 mM Imidazole)
Lane 7: Elution (200 mM Imidazole)
Lane 8: Elution (300 mM Imidazole)
Lane 9: Elution (500 mM Imidazole)
Packing: Ni-IDA Agarose Gel FF (Gravity Prepacked Column) (Cat: abs91007-G), 1 mL
Sample: his6-tagged recombinant Streptococcus Protein G produced in E. coli
Binding buffer: 20 mM PB, 150 mM NaCl, pH 7.4
Elution buffer: 20 mM PB, 150 mM NaCl, 20-500 mM Imidazole, pH 7.4
Figure 4 Packing material: Ni-NTA agarose gel FF (medium-pressure prepacked column) (Cat: abs91008-M), 1 mL
Sample: his6-tagged recombinant Streptococcus Protein G produced in E. coli
Binding buffer: 20 mM PB, 150 mM NaCl, pH 7.4
Elution buffer: 20 mM PB, 150 mM NaCl, 20-500 mM Imidazole, pH 7.4
Table Resistance of NTA Series Metal Chelating Fillers
|
type |
Ni-NTA agarose gel FF |
|
Chelating agent 1 mM EDTA |
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Chelating agent 2 mM EDTA |
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Chelating agent 5 mM EDTA |
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Chelating agent 10 mM EDTA |
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Reducing agent 20 mMDTT |
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Reducing agent 20 mM BME |
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Reducing agent 5 mM TCEP |
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Reducing agent 20 mM TCEP |
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Reducing agent 10 mM GSH |
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Denaturant 8 M urea |
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Denaturant 6 M guanidine hydrochloride |
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Detergent 2% Trilaton X-100 |
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Detergent 2% Tween 20 |
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Detergent 2% Tween 80 |
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Base 1 M sodium hydroxide |
Fourth, the product information of Absin tagged protein purification resin
|
Catalog number |
Product name |
specification |
|
abs91007-F |
Ni-IDA agarose gel FF |
10mL/100mL |
|
abs91007-M |
Ni-IDA agarose gel FF (medium pressure prepacked columns) |
1mL/5mL |
|
abs91007-G |
Ni-IDA agarose gel FF (gravity prepacked columns) |
1mL/5mL |
|
abs91008-F |
Ni-NTA agarose gel FF |
10mL/100mL |
|
abs91008-M |
Ni-NTA agarose gel FF (medium pressure prepacked columns) |
1mL/5mL |
|
abs91008-G |
Ni-NTA agarose gel FF (gravity prepacked columns) |
1mL/5mL |
Absin provides antibodies, proteins, ELISA kits, cell culture, detection kits, and other research reagents. If you have any product needs, please contact us.
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Absin Bioscience Inc. |
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