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Protein Expression & Purification

Products >  Protein_Expression_and_Purification >  His-Tagged_Protein_Purification >  Cobalt_Resin-Cell_Lysate

His-Tagged Protein Purification from Crude Cell Lysates—TALON CellThru

TALON CellThru is a novel IMAC resin for purifying his-tagged proteins from crude cell lysates, sonicates, and fermentation harvests. This resin uses the same proprietary ligand as our TALON Metal Affinity Resin, but has larger beads (300–500 µm) which permit cellular debris to flow through without centrifugation. TALON CellThru Resin captures his-tagged protein directly from crude lysates in one quick step, minimizing protein degradation and generating higher yields of purified protein than conventional strategies.

TALON CellThru Resin is ideal for purifying membrane-bound proteins or multi-protein complexes, which typically pellet with the cell debris during the lysate clarification step and so are not available for binding to the resin. Because crude, nonclarified lysates can be applied directly to the resin, you can skip the centrifugation step, allowing membrane-bound proteins and multi-protein complexes to remain in the lysate where they can bind to the resin (see example).

The resin consists of large agarose beads that can be packed into standard chromatography columns with large-pore frits to prevent column blockage. Particulate material flows between the beads while the soluble protein product is captured by the ion-exchange functional groups attached to the beads. The resin is ideal for use in expanded bed chromatography for recovering even higher amounts of protein and is recommended for applications such as high-throughput purification of his-tagged proteins from crude extracts. We also offer CellThru 10 ml Disposable Columns (with a filter pore size of 90–130 µm) that allow cellular debris to flow through easily.

Choice of Native or Denaturing Purification Conditions

TALON Resin retains its protein binding specificity and yield under a variety of purification conditions. It is stable under both denaturing and native (nondenaturing) conditions (see flow chart). Deciding whether to use native or denaturing purification conditions depends on protein location, solubility, accessibility of the his tag, downstream applications, and preservation of biological activity.

  • Native Conditions

    Purifying a protein under native conditions (see example) is the most efficient way to preserve its biological activity, but requires that the protein be soluble. Advantages include:

    • Eliminating the renaturation step at the end of the purification, saving time, and preventing significant loss of activity
    • Retaining the ability to copurify enzyme subunits, cofactors, and associated proteins
  • Denaturing Conditions

    Because proteins that are overexpressed in prokaryotic systems sometimes form insoluble aggregates called inclusion bodies, you may need to purify proteins under denaturing conditions (see example)—using strong denaturants such as 6 M guanidinium or 8 M urea to enhance protein solubility. Advantages include:

    • Complete solubilization of inclusion bodies and his-tagged proteins
    • Improved binding to the matrix and reduced nonspecific binding, due to full exposure of the his tag

His-tagged proteins purified under denaturing conditions can be used directly in subsequent applications, or may need to be renatured and refolded. Protein renaturation and refolding can be performed prior to elution from the column. However, yields of recombinant proteins will be lower than under native conditions, because urea and guanidinium molecules compete with histidines for binding to metal.

Use of Reducing Agents

Purification with TALON Resin may be carried out in the presence of β-mercaptoethanol (see example), but not DTT or DTE, to preserve reduced sulfhydryl (-SH) groups that are important for the biological activity and structure of a given protein. TALON provides higher yields than Ni-NTA (see example) in the presence of β-mercaptoethanol.

  At-A-Glance   Documents   Images & Data   Resources


  • Purification from crude cell lysate
  • Ideal for purification of membrane-bound protein or multiprotein complexes
  • No copurification of proteins
  • Low metal ion leakage
  • Fast & easy protocol


Purification of protein from:

  • Crude cell lysates
  • Sonicates
  • Fermentation harvests

Reagents Compatible with TALON Resin

Reagent Acceptable Concentration
Beta-mercaptoethanola 10 mM (with caution)
CHAPSb 1% (with caution)
Ethanolc 30%
Ethylene glycol 30%
Glycerol 20%
Guanidine hydrochloridea 6 mM
Imidazoled 200 mM at pH 7.0–8.0, for elution
KCl 500 mM
MES 20 mM
MOPS 50 mM
NaCl 1.0 M
NP-40 1%
SDSb 1% with caution
Trise 50 mM
Triton-X 100 <1%
Urea 8 M

a Use resin immediately after equilibrating with buffers containing these reagents. Otherwise, the resin will change color. Do not store resin in buffers containing these reagents.
b 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.
c Ethanol may precipitate proteins, causing low yields and column clogging.
d Imidazole cannot be used at concentrations higher than 5–10 mM for loading his-tagged proteins, because it competes with the histidine side chains (imidazole groups) for binding to the immobilized metal ions.
e Tris coordinates weakly with metal ions, causing a decrease in capacity.

Reagents Incompatible with TALON Resin

These reagents are incompatible at any concentration:

  • DTT (dithiothreitol) and DTE (dithioerythritol)
    NOTE: Using strong reducing agents will interfere with cobalt metal ion binding to the resin.

  • EDTA (ethylenediaminetetraacetic acid), EGTA (ethylene glycol tetraacetic acid), and PEI
    NOTE: Although you can use EDTA at indicated points, it must be removed from the sample by gel filtration prior to applying the sample to TALON Resins.

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 License Units Select
635513 CellThru 10-ml Disposable Columns 20 Columns $104.00  
635509 TALON® CellThru 10 mL $114.00 License Statements
635510 TALON® CellThru 100 mL $809.00 License Statements


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