Ni-NTA-Nanogold® is designed for detection or localization of polyhistidine (his) -tagged fusion proteins using electron microscopy, light microscopy or blotting.
Ni-NTA-Nanogold® comprises a 1.8 nm or 5 nm Nanogold® particle with multiple nickel-nitrilotriacetic acid functionalities incorporated into the ligands on the surface of gold particles. Nickel-nitrilotriacetic acid functionalities bind to histidines from the tagged proteins, and form stable complexes with extremely low dissociation constants.
Using Ni-NTA-Nanogold®, his-tagged fusion proteins originating from any of a variety of expression vectors can be labeled under non-denaturing or denaturing conditions. The labeled his-tagged fusion proteins can be visualized by microscope or blots, when used with gold or silver enhancement reagents such as our GoldEnhance™, or HQ Silver.
Hybrid method to elucidate protein architecture with nanometer precision:
Developed by Dr. Wei-hau Chang's group (Chang, 2013)
Use 1.8 nm and 5 nm Ni-NTA-Nanogold® together in cryo-EM
for better single particle analysis
Combine with FRET to further refine your protein map
and observe conformational/postioning changes during interactions [ LEARN MORE ]
Nickel staining His-tags for EM: High-resolution labeling of proteins, protein complexes or organelles containing recombinant His-tagged proteins for TEM or STEM localization.
"Universal" pre-embedding labeling of His-tagged proteins in tissue sections for electron microscopic observation.
identifying His-tagged proteins in fractions during Ni-NTA-column purifications.
Detection of recombinant His-tagged proteins on blots and in gels.
Heavy atom labeling of regular structures for image analysis and structure solution.
Optimum size for direct EM viualization Use for post-embedding labeling
Our new 10 nm Ni-NTA-Nanogold® brings optimum EM visualization without silver or gold enhancement, while preserving the high resolution of the original 1.8 nm and 5 nm probes.
The entire probe is similar in size to an unlabeled IgG molecule, but the larger gold particles may be clearly visualized by standard TEM without silver or gold enhancement, even in wider views such as thick sections and whole cells. Blot sensitivity will be even higher that with the smaller sizes due to the larger gold.
High visibility: the 10 nm gold particle is highly monodisperse and clearly visualized at TEM resolution in most kinds of specimens without silver or gold enhancement.
Universal probe for labeling any His-tagged fusion proteins: this simplifies labeling as it may be used to label many different targets without the need for generating different antibodies. It may also be used for multiple labeling studies in conjunction with gold particles of different sizes, or with silver or gold-enhanced Nanogold®.
Precise labeling resolution: the link between the nitrilotriacetic acid - Ni(II) chelate and gold surface is much smaller than an antibody or protein, and therefore bound gold is located much closer to its target: we estimate that the distance from the gold particle surface to the His tag is on the order of 2 nm. Plus, this is a primary probe: there is no secondary antibody to increase the distance to the target. Use NTA-Ni(II)-Nanogold® to localize sites in protein complexes or other macromolecular assemblies at molecular resolution.
High solubility, biocompatibility, and stability:10 nm Ni-NTA-Nanogold® is prepared using a stable, highly hydrophilic surface functionalization.
Strong binding: binding constants for Ni(II)-NTA are very high due to the combination of the chelate effect of multiple histidine binding, and target binding of multiple Ni(II)-NTA functionalities. Dissociation constants are estimated to be between 10-7 to 10-13 M-1. For many applications, this provides binding strengths comparable to antibodies.
High penetration: 10 nm Ni-NTA-Gold is similar in size to an unlabeled antibody IgG molecule, and much smaller than a 10 nm colloidal gold-antibody conjugate. It can therefore penetrate into specimens and access sterically restricted interior sites much better than a 10 nm immunogold probe, without the additional mass of an unlabeled primary, and provide higher labeling density. In some systems it may be used with stronger fixation or less permeabilization, enabling labeling with better ultrastructural preservation.
Super sensitivity: the larger gold particle provides highest sensitivity with virtually no background when used to detect His-tagged targets on blots.
Top: Structure of NTA-Ni(II)-5 / 10 nm Nanogold®, showing the binding of the incorporated metal chelate to a His-tagged protein. Distance from the gold particle surface to the His tag is estimated to be 1.5 nm. Above: Transmission electron micrograph of 5 nm NTA Nanogold: average diameter 5.11±0.84nm.
5 nm Ni-NTA-Nanogold®
Go straight to EM- no silver or gold enhancement needed!
5 nm Ni-NTA-Nanogold® provides new features, improved performance, and extends NTA-Ni(II) targeting technology to larger gold.
This probe has the same high resolution as the 1.8 nm Ni-NTA-Nanogold®. The entire probe is still smaller than an IgG molecule, but the larger gold particles may be clearly visualized by standard TEM without silver or gold enhancement, even in wider views such as thick sections and whole cells. Blot sensitivity will also be higher due to the larger gold.
High visibility: The 5 nm gold particle may be clearly visualized at TEM resolution in wder, cellular views without the need for silver or gold enhancement.
Universal probe for labeling any His-tagged fusion proteins:This simplifies labeling as it may be used to label many different targets without the need for generating different antibodies. It may also be used for multiple labeling studies in conjunction with gold particles of different sizes, or with silver or gold-enhanced Nanogold.
Precise labeling resolution: the nitrilotriacetic acid - Ni(II) chelate is much smaller than an antibody or protein, and therefore when it is bound, the gold is much closer to its target: we estimate that the distance from the gold particle surface to the His tag is on the order of 1.5 nm. This makes NTA-Ni(II)-Nanogold® ideal for localizing sites in protein complexes or other macromolecular assemblies at molecular resolution.
High solubility, biocompatibility and stability: 5 nm Ni-NTA-Gold is prepared using a stable, highly hydrophilic surface functionalization.
Strong binding: binding constants for Ni(II)-NTA are very high due to the combination of the chelate effect of multiple histidine binding, and target binding of multiple Ni(II)-NTA functionalization. Dissociation constants are estimated to be between 10-7 to 10-13 M-1. For many applications, this provides binding strengths comparable to antibodies.
High penetration: 5 nm Ni-NTA-Nanogold® is smaller than an unlabeled primary antibody, can more easily penetrate into specimens and access sterically restricted interior sites, and perturbs the ultrastructure less. In some systems it may be used with stronger fixation or less permeabilization, enabling labeling with better ultrastructural preservation.
Super sensitivity: the larger gold particle provides higher sensitivity with virtually no background when used to detect His-tagged targets on blots. 10 ng of His-tagged ATF-1 was detected without silver or gold enhancement. Gold enhancement allows the detection of 0.5 ng, with no visible binding to an E Coli. extract control.
Versatile: detects and localizes His-tagged recombinant proteins in electron microscope, light microscope and blots.
Stable and soluble: surface functionalization and shell confer high stability combined with high solubility, making Ni-NTA-Nanogold® compatible with most systems.
High resolution: 1.8 nm diameter gold particle provides high resolution for accurate, precise molecular and macromolecular identification and ultrastructural studies on isolated protein complexes and macromolecules.
Permanent: stain does not fade.
left: Structure of Ni-NTA-Nanogold® showing interaction with Interaction with a His-tagged protein; right: Knob protein from adenovirus cloned with 6x-His tag, labeled with Ni-NTA-Nanogold, column purified from excess gold, and viewed in the scanning transmission electron microscope (STEM) unstained (Full width approximately 245 nm).
Custom conjugation is also available, of Nanogold®, FluoroNanogold, undecagold
or colloidal gold to primary antibodies, peptides, small molecules, or other molecules.
International orders:Our regional distributors can provide expedited customs processing and delivery.
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Instructions (PDF)![[Ni-NTA-5 and 10 nm Gold: structure and TEM View (141k)]](../Images/Vol10_Iss7_Fig1_10nm.jpg)
