NTA Agarose for His-tagged Recombinant Proteins

Ni²⁺, Co²⁺, Cu²⁺, or Zn²⁺ charged nitrilotriacetic acid (NTA) coupled to Agarose CL-6B via a stable and uncharged long ether hydrophilic spacer arm, and offers high binding capacity and minimal non-specific binding.

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Description

FastSep high performance resins and chromatography cartridges

Numerous ligand-metal ion combinations
High affinity & high capacity
Minimal non-specific binding & excellent purity
Long spacer arms & minimal steric hindrance
Low metal ion leaching & excellent stability
Maintain high performance after multiple repeated use

Application

For His-tagged protein purification
Simply replace your existing NTA resin or cartridge, no optimization or protocol changes necessary
Purification under native and denaturing conditions
Suitable for small proteins, large protein complexes, and proteins with low expression rates

Purification of His-tagged Recombinant Proteins

A his-tag, or polyhistidine tag, is a string of histidine residues at either the N or C terminus of a recombinant protein. Such a string can have 4 to 10 residues, with 6 being the most common a hexahistidine tag. Recombinant proteins can have more than one hexahistidine tag.

In IMAC, metal ions are immobilized on a resin matrix using a chelating agent such as NTA or IDA.

Ni²⁺, Co²⁺, Cu²⁺ and Zn²⁺are the most commonly used ions for his-tag purification of a recombinant protein. The his-tag has a high affinity for these metal ions and binds strongly to IMAC column, while most other proteins in the lysate will not bind to resin, or bind only weakly. The use of a his-tag and IMAC can therefore provide relatively pure recombinant protein directly from a crude lysate.

How to choose different ligands and metal ions in IMAC?

NTA and IDA are the two most widely used ligands in IMAC. NTA is a tetravalent ligand with coordination number 4 and therefore stronger coordination of metal ions, while IDA is a trivalent ligand with coordination number 3. Generally, NTA gives higher binding specificity and lower metal ion leaching, while IDA shows more non-specific binding and increased metal ion leaching. IDA can have higher metal ion loading density and usually requires much lower imidazole concentration in eluent.

For metal ions going from Co2+, Zn2+, Ni2+ to Cu2+, generally binding affinity increases while binding specificity decreases. Cu2+ has the highest affinity giving the highest protein recovery, but low specificity. Co2+ has the lowest affinity but highest specificity, reducing the amount of impurity proteins in eluate. Ni2+ is often most widely used because it provides a good balance between affinity and specificity.

As many factors can affect the choice of optimal ligand-metal ion combination, the best approach is to try different combinations and let your proteins decide. FastSep(TM) high performance resins and cartridges offer numerous ligand-metal ion combinations to meet various purification needs.
FastSep(TM) high performance resins and chromatography cartridges are based on highly cross-linked agarose 6% beads which have structural characteristics that result in superb physical strength, excellent flow properties, low backpressure, and an open pore structure. FastSep(TM) high performance resins and chromatography cartridges allow operation at very high flow rates, resulting in increased throughput and rapid column cleaning between chromatography runs.

Name	Cat#	Description (every combination is 50% suspension)
Ni-NTA Resin	NINTA-25	25ml nickel-charged NTA resin (50ml total volume)
Ni-NTA Resin	NINTA-100	100ml nickel-charged NTA resin (200ml total volume)
Ni-NTA Resin	NINTA-500	500ml nickel-charged NTA resin (1000ml total volume)
Co-NTA Resin	CONTA-25	25ml cobalt-charged NTA resin (50ml total volume)
Co-NTA Resin	CONTA-100	100ml cobalt-charged NTA resin (200ml total volume)
Co-NTA Resin	CONTA-500	500ml cobalt-charged NTA resin (1000ml total volume)
Cu-NTA Resin	CUNTA-25	25ml copper-charged NTA resin (50ml total volume)
Cu-NTA Resin	CUNTA-100	100ml copper-charged NTA resin (200ml total volume)
Cu-NTA Resin	CUNTA-500	500ml copper-charged NTA resin (1000ml total volume)
Zn-NTA Resin	ZNNTA-25	25ml zinc-charged NTA resin (50ml total volume)
Zn-NTA Resin	ZNNTA-100	100ml zinc-charged NTA resin (200ml total volume)
Zn-NTA Resin	ZNNTA-500	500ml zinc-charged NTA resin (1000ml total volume)
XX-NTA Resin	XXNTA-OEM	Any size any metal ion charged NTA resin

Specification

Chromatography technique	Histidine-tagged protein purification
Matrix	Highly cross-linked 6% beaded agarose
Active group	Ni²⁺, Co²⁺, Cu²⁺, or Zn²⁺ charged Nitrilotriacetic Acid (NTA)
Active group density	>40 umol/ml drained medium
Binding capacity	>40 mg/ml drained medium
Spacer	17 atom stable and uncharged ether hydrophilic linkage
Bead geometry & size	Spherical 50 to 150 um
Bead mean diameter d50v	90 um
Linear flow velocity	<75 cm/h at 25 °C, HR 16/10 column, 5 cm bed height
Recommended linear flow rate	<30 cm/h
Pressure/flow specification	Base matrix 100-200 cm/h, pressure drop cm H2O/bed height = 15, bed height 10 cm, 5 cm i.d.
1ml Cartridge bed dimensions	7×25 mm
1ml Cartridge bed height	25 mm
1ml Cartridge bed volume	1 ml
1ml Cartridge column I.D	7 mm
5ml Cartridge bed dimensions	16×25 mm
5ml Cartridge bed height	25 mm
5ml Cartridge bed volume	5 ml
5ml Cartridge column I.D	16 mm
Maximum cartridge flow rate	5 ml/min (1 ml cartridge) or 20 ml/min (5ml cartridge)
Recommended cartridge flow rate	1 ml/min (1 ml cartridge) or 5 ml/min (5 ml cartridge)
Maximum pressure during operation	5 bar [0.5 MPa] (70 psi)
pH Stability working range	3 to 13
pH Stability cleaning in place (cip)	2 to 14
Chemical stability	Stable to commonly used aqueous solutions. Can be used with non-ionic detergents, denaturing solvents, e.g. 8 M urea and 6 M guanidine hydrochloride, Stable in organic solvents, e.g. 50% methylformamide and 50% dioxane.
Storage	2 to 8 °C