6xhis Tag
6xhis Tag - Related Videos
11:26
High-throughput Gene Tagging in Trypanosoma brucei
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Cited by 21 •
2016
Addition of a tag to a protein is a powerful way of gaining insight into its function. Here, we describe a protocol to endogenously tag hundreds of Trypanosoma brucei proteins in parallel such that genome scale tagging is achievable.
10:32
Orthogonal Protein Purification Facilitated by a Small Bispecific Affinity Tag
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Cited by 7 •
2012
A novel and highly efficient two-step affinity chromatography protocol has been developed and is described in detail. The method is based on a small purification tag with two inherent affinities and is applicable to a wide range of target proteins with different properties.
01:24
Tagging and Fusion Proteins
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2023
Proteins are involved in several cellular processes and biochemical reactions. Analyzing a specific protein of interest requires it to be isolated from the other proteins in the cell. This is achieved by overexpressing the specific gene in a suitable host to produce large quantities of the target protein. A tag or label is recombined with the gene to produce a fusion protein containing the target protein and the tag. The tags on these fusion proteins can then be used for easy detection and...
04:28
A Procedure for the Purification of a Polyhistidine-Tagged Protein from Streptococcus mutans
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2025
Source: Murata, T., et al. Purification of a High Molecular Mass Protein in Streptococcus mutans. J. Vis. Exp. (2019)This video demonstrates a step-by-step procedure for the purification of a polyhistidine-tagged protein secreted from Streptococcus mutans.
07:44
High-throughput Purification of Affinity-tagged Recombinant Proteins
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Cited by 1 •
2012
We describe a method for the affinity-tagged purification of recombinant proteins using liquid-handling robotics. This method is generally applicable to the small-scale purification of soluble His-tagged proteins in a high-throughput format.
FAQs
Binding depends on coordination between histidine imidazole groups and immobilized metal ions. This interaction gives the tagged protein affinity for the resin, while many untagged cellular proteins do not remain bound during washing. The separation therefore relies on differential retention, allowing the target protein to be enriched before release.
Nickel and cobalt provide alternative metal ions for the affinity resin. Once the tagged protein is retained, imidazole or an altered pH can disrupt the interaction and release it. This two-stage behavior separates capture from recovery, so the purification outcome depends on retaining binding until elution.
The small size of the 6xHis tag supports streamlined workflows for examining recombinant proteins. In practice, the same fusion can facilitate isolation for structural or functional studies and support analysis of protein interactions or activity. Its value is therefore not limited to purification; it can connect sample preparation with downstream biological characterization.
A typical workflow begins with expression of a recombinant protein fused to the tag, followed by loading the sample onto a metal-affinity resin. Washing removes untagged cellular proteins, and imidazole or altered pH releases the retained target. These linked steps produce an enriched protein preparation for subsequent detection or studies of structure, function, interactions, or activity.
Beyond purification, 6xHis tagging supports detection of recombinant proteins through immunodetection. This gives researchers a way to analyze whether the tagged protein is present after expression or sample processing. Used alongside affinity purification, detection connects recovery of the target with its subsequent biological analysis in the same experimental workflow.
In biology, the tag links recombinant protein expression to purification and analysis within one experimental strategy. A researcher can use affinity capture to obtain an enriched target and then examine it through immunodetection or studies of structure, function, interactions, or activity. This integration helps organize workflows around the same engineered protein sample and its downstream characterization.