The polyhistidine tag / Poly-His-tag is the most widely used protein affinity tag. Due to its small size of only six amino acids it rarely interferes with the protein's function. Additionally its low immunogenicity and versatility under native and denaturing conditions are highlighting this protein tag among others. The underlying purification principle of the His-tag is an interaction between metal ions and the imidazole ring of histidine. Further information to His-tags
can be found here.
Cube Biotech's His tag purification products:
To know about His-tag purification resins and magnetic beads:
Choosing the correct type of His-tag purification resin or magnetic bead is based on three factors:
- The Ligand: This is the molecule to which the metal ion is coupled in a chelator complex (see Figure 1).
Cube Biotech offers three different ligands.
- NTA: Nitrilotriacetic acid is the most commonly used ligand. It's chelator complex with Nickel is depited in figure 1.
- IDA: Iminodiacetic acid is the second most commonly used one. Click here to learn the differences between IDA and NTA.
- INDIGO:This ligand was developed here at Cube Biotech. Among its advantages is a highly increased DTT and EDTA tolerance. Click here to learn more about INDIGO.
- The coupled metal Ion: Depending on which metal ion is coupled to the ligand the affinity and specificty of the His-tag purification changes. Rule of thumb: The higher the protein yield, the less specific the purification gets. Figure 2 gives an overlook one the affinty/specificty ratio of the most commonly used metal ions for His-tag purification.
- The size of the beads: The bigger the used beads are in diameter the higher the flow rate for FPLC purification. However having a slower flow rate usually increases the protein yield. We offer bead diameters of 30 µm, 40µm, 100 µm and XL sized (~400 µm). Further sizes on request available. See figure 3 for reference.
His-tag affinity resins and MagBeads in detail:
Figure 1: Schematic depiction of a chelator complex conisting of NTA and a Nickel2+
. This Nickel Ion binds to two imidazol rings of a His-tag that has been added to a protein.
Figure 2: Overview on the most commonly used metal ions for His-tag protein purification. Copper has the highest affinity, while in turn having the lowest specificity. Cobalt marks the other end of the spectrum. Nickel, the most commonly used metal tions has a good balance between affinity and specificity.
Figure 3: Our commonly provided bead sizes in compairson. The smallest beads are magnetic beads with a 30 µm diameter and agarose resin with 40 µm. Our XL sized beads with 400 to 500 µm diameter are marking the other end of the spectrum. Sizes above that are depicted but have to be ordered on request.