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
Ligands	Iminodiacetic acid, ~30μmol Ni2+/mL	Hypotritriacetic acid, ~17μmol Ni2+/mL
Particle size rangea	45~165μm
Average particle size	~90μm
Dynamic binding capacityb	≥ 40 mg/mL histidine-tagged protein
Recommended workflow speed	60~300cm/h
Maximum flow rate and pressure	＞1500cm/h，0.5MPa
Use pH	4~8.5 (recommended working pH), 3~12 (long-term stability); 2~14(Short-term stability)
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

 

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
Chelating agent 2 mM EDTA
Chelating agent 5 mM EDTA
Chelating agent 10 mM EDTA
Reducing agent 20 mMDTT
Reducing agent 20 mM BME
Reducing agent 5 mM TCEP
Reducing agent 20 mM TCEP
Reducing agent 10 mM GSH
Denaturant 8 M urea
Denaturant 6 M guanidine hydrochloride
Detergent 2% Trilaton X-100
Detergent 2% Tween 20
Detergent 2% Tween 80
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.

Absin Bioscience Inc. Email: worldwide@absin.net

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