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Protein Expression & Purification

Products >  Protein_Expression_and_Purification >  His-Tagged_Protein_Purification >  Nickel_Columns-FPLC

His60 Ni Superflow Cartridges Using FPLC

His 60 Ni Superflow, the highest-capacity resin to date that provides great purity, is now available in prefilled 1 ml and 5 ml cartridges for his-tagged protein purification on any LC system, such as ÄKTA or FPLC, or for manual purification using a syringe. The resin enables fast, easy, and reproducible chromatographic separations and can be regenerated for multiple uses.

Protein Purification Principle

His60 Ni Protein Purification Resin, which is based on a modified Ni-IDA resin, provides remarkable selectivity and reliability when is it used to purify proteins containing an affinity tag of six or more consecutive histidine residues—such as the 6xHis tag or other his-tags. This technology allows one-step purification of almost any his-tagged protein from any expression system, under native or denaturing conditions. The His60 Ni Superflow purification resin has low nickel ion leakage and the highest binding capacity to date (outperforming Ni-NTA and other resins).

His-Tagged Protein Purification Procedure

The purification of recombinant his-tagged proteins consists of 4 steps: cell lysis/extraction, binding, washing, and elution. His60 Nickel allows one-step protein purification under either native or denaturing conditions. The resin is compatible with multiple denaturants and detergents. Please see user manual for complete reagent compatibility list. Purified recombinant his-tagged proteins can be eluted by the addition of 300 mM imidazole or by a reduction in pH. To protect your his-tag protein from degradation it is recommended to use protease inhibitors (ProteoGuard) during the purification process.

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  • Up to 60 mg/ml of recombinant his-tagged protein (1 ml cartridge)
  • Up to 300 mg/ml of recombinant his-tagged protein (5 ml cartridge)
  • Low metal ion leakage in eluted his-tagged protein
  • For purification under native or denaturing conditions
  • Fast and reproducible processing on any LC system, including ÄKTA


Purified recombinant his-tagged proteins can be used for:

  • Protein labeling
  • Immunization to produce antibodies
  • Animal studies
  • Other applications

His60 Ni Superflow Cartridge Specifications

1 ml Cartridge

5 ml Cartridge


Superflow 6 (cross-linked agarose)

Bead Diameter

60–160 µm

Column Dimensions (mm i.d.)

0.7 cm x 2.5 cm

1.6 cm x 2.5 cm

Maximum Pressure1

5 bar, 0.5 MPa

Typical Back Pressure

1.0 bar, 0.1 MPa (1 ml/min)

2.0 bar, 0.2 MPa (5 ml/min)

Recommended Flow Rate

1 ml/min (156 cm/hr)

5 ml/min (149 cm/hr)

Maximum Flow Rate2

10 ml/min (1,559 cm/hr)

40 ml/min (1,193 cm/hr)

pH Stability Short Term (<2 hr)


pH Stability Long Term (>2 hr)


Binding Capacity3

60 mg of AcGFPuv

300 mg of AcGFPuv

System Compatibity4

Automated chromatography systems
(e.g. ÄKTA, FPLC, etc.), peristaltic pump or syringe

Cartridge Body Material



1/16"(inlet); 1/16" (outlet)

1 The maximum pressure that can be used with the Superflow matrix itself is 10 bars. However, stability of the cartridges is only guaranteed up to 5 bars.
2 High flow rates may lead to reduced recovery of 6xHis-tagged protein.
3 Binding capacity may vary from protein to protein.
4 Adaptors may be necessary.

Additional Products

His60 Ni resin (a nickel-based resin) is sold separately as:   TALON resin (a cobalt-based resin) is sold separately as:Detect his-tagged proteins with his tag antibodies

Reagents Compatible with His60 Ni Superflow Resin

Reagent Acceptable Concentration
Beta-mercaptoethanola 20 mM (with caution)
CaCl2 5 mM
CHAPSb,c 1% (with caution)
DTT (dithiothreitol)d 1 mM (with caution)
Ethanole 20%
Glycerol 20%
Guanidine hydrochloridef 6 mM
HEPESg Up to 100 mM (with caution)
Histidineh 20 mM to inhibit nonspecific binding.
Up to 100 mM to elute his-tagged proteins.
MgCl2 4 M
MOPSg Up to 100 mM (with caution)
NP-40b,i 2%
SDSb,c 1% (with caution)
Sodium acetate Up to 100 mM (with caution)
Sodium phosphate Up to 50 mM
Trisj Up to 50 mM (with caution)
Triton-X 100b,i 1%
Tween 20 2%
Ureaf 8 M

a Use the resin immediately after equilibrating with buffers containing beta-mercaptoethanol. Otherwise, a slight change in color (yellowing of the resin) will occur. Do not store the resin in buffers containing beta-mercaptoethanol.
b Detergents cannot be easily removed by buffer exchange.
c Ionic detergents like CHAPS (3-[(3-Cholamidopropyl)-dimethylammonio]-1-propane-sulfonate), SDS (sodium dodecyl sulfate), and sarkosyl are compatible up to 1%. However, due to their charged nature, you should anticipate interference with binding, even at low concentrations.
Since DTT is a reducing agent, low concentrations will reduce the metal ions in His60 Ni Superflow resin. Although enough of these ions may remain unaffected to allow protein purification, please use it with caution. Do at least 20 column volumes of washes, preferably with low concentrations of imidazole (40 mM) to wash out any reduced metal ions.
e Ethanol may precipitate proteins, causing low yields and column clogging.
f With high concentrations, protein unfolding generally takes place. Protein refolding on-column (or after elution) is protein-dependent.
g Amine groups that are present in these buffers can interact with Ni2+ ions, diminishing the resin’s binding capacity.
h Binds to His60 Ni and competes with histidine residues in the his tag.
i Has high absorbance at 280 nm.
j Tris coordinates weakly with metal ions, causing a decrease in capacity.

Reagents Incompatible with His60 Ni Superflow Resin

These reagents are incompatible at any concentration:

  • Arginine, glycine, and glutamine

  • EDTA (ethylenediaminetetraacetic acid), EGTA (ethylene glycol tetraacetic acid), and PEI
    NOTE: Using these chelating agents will strip metal ions from the resin, resulting in protein elution and a resin color change.

  • DTE (dithioerythritol)

Additional Information

Please see the product's Certificate of Analysis for information about storage conditions, product components, and technical specifications. Please see the Kit Components List to determine kit components. Certificates of Analysis and Kit Components Lists are located under the Documents tab. 

Cat. # Product Contents Size Price Units Select
635680 His60 Ni Superflow Cartridge 1 x 5 mL Cartridge $125.00
635674 His60 Ni Superflow Cartridge Purification Kit 5 x 1 mL Cartridges $420.00
635678 His60 Ni Superflow Cartridge Purification Kit 5 x 5 mL Cartridges $714.00
635675 His60 Ni Superflow Cartridges 5 x 1 mL Cartridges $191.00
635679 His60 Ni Superflow Cartridges 5 x 5 mL Cartridges $500.00


Recommended Products
Cat. # Product Contents Size Price Units Select
631210 6xHis mAb-HRP Conjugate 100 uL $350.00
631212 6xHis Monoclonal Antibody (Albumin Free) 200 ug $509.00
635665 His60 Ni Buffer Set 20 Purifications $253.00
635673 ProteoGuard™ EDTA-Free Protease Inhibitor Cocktail 10 x 100 uL $140.00

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