HisTALON™ Gravity Column Purification Kit, 5 x 1 mL Columns, 635654, Takara Bio Inc (CAT#: STEM-C-3814-LGZ)

TALON Metal Affinity Resin is a durable immobilized metal affinity chromatography (IMAC) resin with remarkable affinity and specificity for his-tagged proteins. TALON resins are compatible with all commonly used IMAC reagents and allow protein purification under native or denaturing conditions.

TALON metal affinity resins use cobalt ions to purify recombinant his-tagged proteins. TALON is a cobalt-containing tetradentate chelator that is specific for his-tagged proteins. The TALON reaction core contains cobalt, which has strict requirements on the spatial positioning of histidine. Only adjacent histidines or specially positioned histidines can combine with cobalt in the reaction core. In nickel-based resins (ie, Ni-NTA resins), these space requirements are less stringent. Therefore, nickel-based resins are also capable of binding histidines located at positions other than the his-tag of proteins.

Takara Bio offers ready-to-use HisTALON gravity columns for efficient purification of HisTALON-tagged proteins from bacterial, mammalian, and baculovirus-infected cells using a gravity-flow-based protocol. The column is pre-packed with our TALON resin and can absorb more than 20 mg of TALON-tagged AcGFP1. These columns enable fast, easy, reproducible chromatographic separations and can be regenerated for a variety of uses. However, we recommend that you only reuse columns when purifying different batches of the same protein. If you plan to purify multiple proteins on the same column, you must use the "full regeneration" method described in the user manual.

Choice of native or denaturing purification conditions

TALON resin maintains its protein binding specificity and yield under a variety of purification conditions. It is stable under both denaturing and native (non-denaturing) conditions. The decision to use native or denaturing purification conditions depends on the protein's location, solubility, availability of his-tags, downstream applications, and the need to preserve biological activity.

Purifying proteins under native conditions is the most effective way to maintain their biological activity, but requires the protein to be soluble. Advantages include:

Eliminates the redenaturation step at the end of purification, saving time and preventing significant loss of activity.
Retains the ability to co-purify enzyme subunits, cofactors, and associated proteins.

Because proteins overexpressed in prokaryotic systems sometimes form insoluble aggregates called inclusion bodies, you may need to purify the protein under denaturing conditions - use strong denaturing agents such as 6M guanidine or 8M urea to enhance protein solubility. Advantages include:

Complete dissolution of inclusion bodies and his-tagged proteins.
Improved binding to matrix and reduced non-specific binding due to fully exposed his-tag.

His-tagged proteins purified under denaturing conditions can be used directly for subsequent applications, or may require renaturation and refolding. Protein refolding and refolding can be performed prior to elution from the column. However, because urea and guanidine molecules compete with histidine for metal binding, the yield of recombinant protein will be lower than under native conditions.

Use of reducing agent

Purification of TALON resins can be performed in the presence of β-mercaptoethanol instead of DTT or DTE to preserve reduced sulfhydryl (-SH) groups that are important for the biological activity and structure of specific proteins. In the presence of β-mercaptoethanol, the yield of TALON was higher than that of Ni-NTA.

Size: 5 x 1 mL Columns

Easy-to-use prepacked gravity columns
Maximize yield of biologically active protein
Superior purity & high yields with TALON Resin

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