Ni-NTA Superflow Cartridges

For His-tagged protein purification using liquid chromatography systems

✓ 24/7 automatic processing of online orders

✓ Knowledgeable and professional Product & Technical Support

✓ Fast and reliable (re)-ordering

Ni-NTA Superflow Cartridges (5 x 5 ml)

Cat. No. / ID: 30761

5 cartridges pre-filled with 5 ml Ni-NTA Superflow: for automated purification of His-tagged proteins using liquid chromatography systems

€867.00

Quantity

Ni-NTA Superflow Cartridges are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.

✓ 24/7 automatic processing of online orders

✓ Knowledgeable and professional Product & Technical Support

✓ Fast and reliable (re)-ordering

Features

Most robust one-step purification over the widest range of conditions
High yields of high-purity protein, with up to 50 mg/ml
Fast, easy, and reproducible processing on any LC system

Product Details

Ni-NTA Superflow, the most-cited resin used for purification of His-tagged proteins, is available in pre-filled 1 ml and 5 ml cartridges for automated purification on liquid chromatography systems such as the FPLC, ÄkTA, and BioLogic systems, or manual purification using a syringe.

Performance

Ni-NTA gives superior performance in delivering high yields of highly purified protein, in comparison to other resins (see figure Ni-NTA outperforms other resins to deliver high yields of high-purity proteins). An independent comparison with other commercially available nickel resins demonstrated that Ni-NTA Superflow shows the lowest level of nickel leaching (see figure An independent study shows that Ni-NTA loses less nickel than any other tested resin). The significance here is that if nickel is leached from the resin, the remaining charged ligands act as an ion exchanger and bind non-tagged proteins that will contaminate elution fractions. The higher stability of Ni-NTA means that even in the presence of 10 mM DTT it can be used to obtain fully active, high-purity proteins (see figure The extra coordination site in Ni-NTA binds nickel ion more tightly than IDA).

As shown in the table, purities are consistently high over all purification scales. Ni-NTA Superflow Cartridges form part of the comprehensive and complementary solutions for His-tagged protein offered by QIAGEN (see figure Scalable purification of nanogram to gram amounts of His-tagged proteins).

Micro- to large-scale purification of IL-1β for a biopharmaceutical project
Matrix	Matrix volume	Culture volume	Yield	Recovery (%)	Purity*
Ni-NTA Magnetic Agarose Beads (micro-scale)	100 μl	1 ml	33 μg	~90%	~97%
Ni-NTA Superflow (small-scale)	500 μl	320 ml	6 mg	~80%	~96%
Ni-NTA Superflow (medium-scale)	10 ml	1.7 l	109 mg	~80%	~98%
Ni-NTA Superflow (large-scale)	100 ml	18 l	2 g	>88%	~97%

* Determined using Agilent Bioanalyzer (Protein 50 LabChip Kit)

See figures

Ni-NTA outperforms other resins to deliver high yields of high-purity protein.

An independent study shows that Ni-NTA loses less nickel than any other tested resin.

The extra coordination site (arrowed) in Ni-NTA binds nickel more tightly than IDA (the ligand used in many competitor resins).

Scalable purification of nanogram to gram amounts of His-tagged proteins.

Principle

Ni-NTA matrices are the affinity chromatography solution of choice for purifying His-tagged proteins (see figure Efficient one-step purification of His-tagged proteins). Their high stability means that they are compatible with a wide range of buffer components, including strong denaturants, detergents, and even reducing agents (see table Reagents compatible with the His/Ni-NTA interaction). This flexibility enables researchers to develop optimal purification schemes while still benefiting from the excellent separation characteristics delivered by Ni-NTA, often making a second chromatographic step unnecessary.

Reagents compatible with the His/Ni-NTA interaction.
Denaturants	Detergents	Reducing agents	Others	Salts	Long-term storage
6 M Gu·HCl	2% Triton X-100	20 mM β-ME	50% glycerol	4 M MgCl₂	Up to 30% ethanol
8 M urea	2% Tween 20	10 mM DTT	20% ethanol	5 mM CaCl₂	or 100 mM NaOH
	1% CHAPS	20 mM TCEP 20 mM	20 mM TCEP	20 mM imidazole	2 M NaCl

See figures

Efficient one-step purification of His-tagged proteins using buffers containing a wide variety of additives.

Procedure

The robust Superflow matrix allows flow rates up to 10 ml/min (1 ml cartridges) and 40 ml/min (5 ml cartridges), speeding the purification procedure and increasing throughput. The cartridges are quickly and easily connected to liquid chromatography systems or a syringe for manual purification. The purification process is highly reproducible, guaranteeing the same high quality protein preparations time after time (see figure ).

See figures

Highly reproducible and reliable purification.

Applications

Ni-NTA matrices can be used to scale up purification of His-tagged proteins for structural studies (e.g., using protein crystallography or NMR), or to produce gram amounts for biopharmaceutical production.

Supporting data and figures

An independent study shows that Ni-NTA loses less nickel than any other tested resin.

An independent study shows that Ni-NTA loses less nickel than any other tested resin. Fifty column volumes of buffer containing various additives was passed through a small column containing 100 μl resin from QIAGEN, Supplier G, S, or I. The flowthrough was pooled and a sample sent for analysis by inductively coupled plasma mass spectrometry (ICP-MS) at Dr.Weßling Laboratories, Bochum, Germany according to DIN EN ISO 17025. Native buffer: 50 mM Na phosphate; 300 mM NaCl; 10 mM imidazole, pH 8.0. Denaturing buffer: 100 mM Na phosphate; 10 mM Tris·Cl; 8 M urea.

Specifications

Features	Specifications
Applications	Proteomics
Gravity flow or spin column	FPLC or syringe
Binding capacity	Up to 50 mg/ml
FPLC	Yes
N- or C-terminal tag	Both
Bead size	60-160 µm
Processing	Manual/Automated
Scale	Large scale
Start material	Cell lysate
Support/matrix	Superflow
Tag	6xHis tag
Yield	Up to 50 mg (1 ml cartridge), up to 250 mg (5 ml cartridge)

Resources

For manual or FPLC purification of His-tagged proteins

Publications

A highly specific system for efficient enzymatic removal of tags from recombinant proteins.

Schäfer F; Schäfer A; Steinert K;

J Biomol Tech; 2002; 13 (3):158-71 2002 Sep PMID:19498979

Use of dual affinity tags for expression and purification of functional peripheral cannabinoid receptor.

Yeliseev A; Zoubak L; Gawrisch K;

Protein Expr Purif; 2006; 53 (1):153-63 2006 Dec 12 PMID:17223358

FAQ

Yes, Ni-NTA and Strep-Tactin Superflow Cartridges can be connected in series.

The Union M6 female/1/16’’ male connector from GE (Code No. 18-3858-01) can be used for this, for example.

FAQ ID -1606

A maximum of 1 hour is needed for the complete purification process with Ni-NTA Superflow Cartridges when recommended flow rates are applied. This is true for 1 ml and 5 ml cartridges.

Details for a typical 1 ml column run:

Equilibration (5 column volumes (cv)): 5 min
Load (10 ml Cleared Lysate): 10 min
Wash (10 cv): 10 min
Elution (10 cv): 10 min
optional: Cleaning-in-place (0.5 M NaOH, 0.5 ml/min, 7.5 cv): 15 min

FAQ ID -1605

The binding capacity of Ni-NTA resins is protein dependent. We guarantee a binding capacity of up to 20 mg/ml of Ni-NTA Superflow for every 6xHis-tagged protein (up to 20 mg per 1 ml Ni-NTA Superflow Cartridge).

However, we have examples where the binding capacity is higher (e.g.: 6xHis-CAT: 30 mg/ml resin; 6xHis-GFP: 55 mg/ml resin).

Binding capacity for the 1 ml Ni-NTA Superflow Cartridge: 20 mg
Binding capacity for the 5 ml Ni-NTA Superflow Cartridge: 100 mg

FAQ ID -1603

If the same protein is purified, the Ni-NTA and Strep-Tactin Superflow Cartridges can be used more than once. Cleaning in place (CIP) is recommended between purifications (0.5 M NaOH, 0.5 ml/min, 7.5 cv for 15 min).

However, for sequential purification of different proteins we recommend to use different cartridges. Please note that cartridges cannot be opened!

FAQ ID -1607