Nanogold®-Antibody and Streptavidin Conjugates

Updated: June 1, 2006

Introduction
Features of Nanogold® Conjugates
Applications:
Custom Labeling
Current Price List

Introduction

Nanogold® is a better gold label. The 1.4 nm Nanogold® particle is a gold compound: it is not just adsorbed to proteins, like colloidal gold, but covalently reacts at specific sites under mild buffer conditions.

The Nanogold® particle is covalently and specifically linked to a hinge thiol on Fab' or IgG. The conjugate therefore has excellent stability compared to colloidal gold-antibody preparations. It is the first probe offered as a Fab' conjugate, which is the smallest gold-antibody probe commercially available. These substantially smaller probes reach more antigens and provide better labeling. They can be viewed directly in TEM without silver enhancement or developed with silver to any appropriate size for enhanced visibility with other counterstains. Whereas the stoichiometry of conventional gold probes particles to IgG molecules varies from 0.2 to 10, Nanogold® preparations contain close to one Nanogold® to one Fab' or IgG.

Features of Nanogold® Conjugates

Smallest commercially available gold immunoprobes.
Penetrates and reaches antigens inaccessible to other probes: proven penetration up to 40 痠 into
Extremely uniform 1.4 nm diameter gold label.
Fab', IgG and Streptavidin conjugates available.
Close to 1 Nanogold® particle to 1 Fab' (or IgG).
Low non-specific affinity gives minimal background.
Ultrasensitivity with silver enhancement: 0.1 pg of antigen detection on immunoblots.
Gold is covalently attached to antibody remote from antigen binding region.
High stability and long shelf life: conjugates show unchanged reactivity after storage for a year.
Stable to a wide range of pH and ionic strengths.

Instructions and technical information (html)
Instructions (PDF)
Material Safety Data Sheet (PDF)

Uses of Nanogold® Conjugates include:

EM of ultrathin cryosections (see: Takizawa, T., and Robinson, J. M.; J. Histochem Cytochem., 42, 1615-1623 (1994)). [Other references].
Pre-embedding electron microscopy. [References]
Post-embedding electron microscopy (for methods for both pre-embedding and post-embedding, see: Nusser, Z., et al.: J. Neuroscience, 15, 2948-2960 (1995)). [Other references]
Correlative electron and light/confocal microscopy (see: Sun, X.J., et al.: J. Histochem. Cytochem., 43, 329-35 (1995)).
In situ hybridization detection. [References]
Scanning electron microscopy (see: Hermann, R., et al.: Histochem. Cell Biol., 106, 31-39 (1996)).

Complete list of references for Nanogold® conjugates
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Applications

In Situ Hybridization

The small size of Nanogold® probes and their consequent ability to access nuclear antigens, combined with the high sensitivity obtained with silver enhancement, makes them excellent reagents for ultrasensitive in situ hybridization detection of specific DNA sequences, both with and without in situ PCR. Striking results have been obtained with amplification of the target signal using biotinylated tyramide (Catalyzed Reporter Deposition or CARD, also known as Tyramide Signal Amplification or TSA): detection of the product with Nanogold®-streptavidin and silver enhancement results in clear signals even in cells with only one or two copies of the target gene (see: Hacker, G. W., et al.: Cell Vision, 4, 54-65 (1997)). Similar sensitivity is seen in RNA detection (see: Zehbe, I., et al.: Am. J. Pathol., 150, 1553-1561 (1997)).

CARD-Nanogold detection of Sinlge-Copy HPV-16 in SiHa cells (59k)

Single copies of HPV 16 in SiHa cells detected by CARD-amplified Nanogold®-silver ISH. This cell line derived from cervical carcinoma contains only one to a few copies of HPV 16, which appear as single spots. Cytospin preparation, counterstained with Nuclear Fast Red (light micrograph courtesy of Dr. G. Hacker, Landeskrankenstalten, Salzbrg. Austria).

Advantages

More sensitive than NBT-BCIP alkaline phosphatase or peroxidase in combination with label amplification for single gene copy resolution.
No need for more lengthy PCR procedures for most cases: avoids false positives due to mispriming and amplicon diffusion.
Black color easily seen with standard brightfield microscope. No expensive fluorescent optics required, and no problems with autofluorescence or bleaching.
Black signal color is compatible with full strength standard cell and nuclear stains (H & E, nuclear fast red, methyl green).
High spatial resolution signal which may be used for EM studies as well.

Special report on In Situ hybridization with Nanogold®

Nanogold®-Streptavidin: instructions and technical information (html)
Instructions (PDF)
Material Safety Data Sheet (PDF)

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Electron Microscopy

STEM Micrograph of Nanogold-IgG conjugate (36k)

This scanning transmission electron microscope (STEM) image clearly shows that labeling with Monomaleimido-Nanogold® (arrows) occurs specifically at a hinge thiol site on the IgG molecule.

Nanogold® sets an unrivaled performance standard for electron microscopy. Nanogold® conjugates diffuse easily tens of microns into tissues, permeabilized cells and nuclei, where most colloidal gold probes have difficulty. Staining is more stoichiometric, with lower backgrounds. Although the 1.4 nm Nanogold® particles are visible directly at 50-120 kX magnification even in 80 nm sections (in otherwise unstained samples) they are more easily visualized after a brief (1-5 min) enhancement with silver developer (LI Silver or HQ Silver). The resulting 2 to 100 nm silver grains (size depends on development time) are easily seen at lower magnification even with uranyl acetate, osmium, or lead citrate staining.

Nanogold®-antibody conjugates Nanogold®-streptavidin

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Light and Confocal Microscopy

Light Micrograph of Nanogold vs. colloidal gold-Labeled Microtubules (81k)

Spindle microtubules labeled with anti-tubulin primary antibody followed by (LEFT) goat anti-mouse colloidal gold or (RIGHT) goat Fab' anti-mouse-Nanogold� (Light micrograph courtesy of Dr. D. Vandré and Dr. R. Burry, Ohio State University. Original magnification = 1300x).

Nanogold® probes have excellent penetration into permeabilized cells and nuclei, similar to immunofluorescent probes and far better than colloidal gold probes. Nanogold®-Fab' probes are smaller than immunofluorescent IgG labels. There is absolutely no aggregation with Nanogold® probes, and they are very stable since the Nanogold® is covalently linked to the conjugate biomolecule. With a subsequent brief (5-15 min) silver enhancement (LI Silver) they give a signal visible even in ordinary brightfield optical microscopes (UV optics are not necessary) with a sensitivity that is usually better than fluorescence. Since the same Nanogold® probe can be used for the EM as well, preliminary staining can be followed in the light or confocal microscope while a parallel sample can be examined at the ultrastructural EM level. This flexibility is not available with fluorescent probes. Nanogold® is now available conjugated to streptavidin and anti-biotin Fab' for nucleic acid detection.

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References for Nanogold® conjugates in light microscopy
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Blotting and Immunochemical Techniques

Nanogold®-antibody conjugates have been used to detect < 0.1 pg (7 X 10^-19 moles) of antigen on immunodot blot assays. This sensitivity is better than most radioactive, enzymatic, colloidal gold, or fluorescence techniques. In fact, Nanogold® labeling is one of the most sensitive assays known. It rivals chemiluminescence, but provides a permanent record without the nuisance of film, development and darkroom procedures, and in a much shorter time. Nanogold® reagents may also be used for Westerns, and our Nanogold®-streptavidin or Nanogold®-anti-biotin conjugates provide ultrasensitive nucleic acid detection.

Immunoblot with Nanogold®

[Blot on the Landscape...] (33k)
Nanogold®-anti-mouse Fab' blotted against mouse IgG, developed with LI Silver (Nanoprobes). These ultra small gold particles nucleate silver deposition so well that unprecedented sensitivity is achieved. This immunodot blot shows 0.1 pg sensitivity (arrow).

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Gel Stain Nanogold®-Labeled Bands

After a protein or other molecule is labeled with Nanogold®, it may be run on a polyacrylamide gel. Subsequently, it may be developed with LI Silver to specifically stain the gold-containing bands. The process is rapid, sensitive and selective, only taking a few minutes for dense staining to develop. It also works on transfer blots. This may be used to distinguish which components are labeled with gold (References)

Ask for application Note 1A "Detection of Nanogold®-labeled molecules on gels." Also see (a) Gregori, L.et al.: J. Biol. Chem., 272, 58-62 (1997), or (b) Wilkens, S., and Capaldi, R. A.: Arch. Biochem. Biophys., 229, 105-109 (1992).

Custom Labeling

custom labeling of commercially available antibodies or your primary antibody with Nanogold® may be possible. Please Call 877-447-6266 (US toll-free number) or (631) 205-9490, use our custom synthesis quotation form, or e-mail us at tech@nanoprobes.com to discuss your project and obtain a quotation.

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To place an order:

Order directly with our online order form

Or contact us:

Price List: Nanogold® Conjugates

Size Price

0.5 mL $182

1.0 mL $291

Instructions and technical information (html)
Instructions (PDF)
Material Safety Data Sheet (PDF)

2001 NANOGOLD®- anti mouse (NMI)
1.4 nm gold particle is attached to affinity-purified IgG molecule,
raised in goat, against mouse IgG (whole molecule).

2002 NANOGOLD®- anti mouse (NMF)
1.4 nm gold particle is attached to affinity-purified Fab' fragment,
raised in goat, against mouse IgG (whole molecule).

2003 NANOGOLD®- anti rabbit (NRI)
1.4 nm gold particle is attached to affinity-purified IgG molecule,
raised in goat, against rabbit IgG (whole molecule).

2004 NANOGOLD® - anti rabbit (NRF)
1.4 nm gold particle is attached to affinity-purified Fab' fragment,
raised in goat, against rabbit IgG (whole molecule).

2005 NANOGOLD®- anti goat (NGI)
1.4 nm gold particle is attached to affinity-purified IgG molecule,
raised in rabbit, against goat IgG (whole molecule).

2006 NANOGOLD®- anti goat (NGF)
1.4 nm gold particle is attached to affinity-purified Fab' fragment,
raised in rabbit, against goat IgG (whole molecule).

2007 NANGOLD- anti rat (NRatI)
1.4 nm gold particle is attached to affintiy-purified IgG molecule,
raised in goat, against rat IgG (whole molecule).

2008 NANOGOLD®- anti rat (NRatF)
1.4 nm gold particle is attached to affinity-purified Fab' fragment,
raised in goat, against rat IgG (whole molecule).

2015 NANOGOLD®- anti biotin (NBI)
1.4 nm gold particle is attached to anti-biotin IgG molecule
raised in goat.

2016 NANOGOLD®- streptavidin (NS)
1.4 nm gold particle is attached to streptavidin.
Instructions and technical information (html)
Instructions (PDF)
Material Safety Data Sheet (PDF)

2050 NANOGOLD®- anti sheep (NSI)
1.4 nm gold particle is attached to affinity-purified IgG molecule,
raised in rabbit, against sheep IgG (whole molecule).

2051 NANOGOLD®- anti sheep (NSF)
1.4 nm gold particle is attached to affinity-purified Fab' fragment,
raised in rabbit, against sheep IgG (whole molecule).

2052 NANOGOLD®- anti human (NHI)
1.4 nm gold particle is attached to affinity-purified IgG molecule,
raised in goat, against human IgG (whole molecule).

2053 NANOGOLD®- anti human (NHF)
1.4 nm gold particle is attached to affinity-purified Fab' fragment,
raised in goat, against human IgG (whole molecule).

2054 NANOGOLD®- anti guinea pig (NGPI)
1.4 nm gold particle is attached to affinity-purified IgG molecule,
raised in goat, against guinea pig IgG (whole molecule).

2055 NANOGOLD®- anti guinea pig (NGPF)
1.4 nm gold particle is attached to affinity-purified Fab' fragment,
raised in goat, against guinea pig IgG (whole molecule).

Instructions and technical information (html)
Instructions (PDF)
Material Safety Data Sheet (PDF)

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Nanogold®-antibody conjugates	Nanogold®-streptavidin
Instructions and Product information (html)	Instructions and product information (html)
Product information (PDF)	Product information (PDF)
Material Safety Data Sheet (PDF)	Material Safety Data Sheet (PDF)

Nanogold®-Antibody and Streptavidin Conjugates

Contents

In Situ Hybridization

Price List: Nanogold® Conjugates