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Updated: July 6, 2001

N A N O P R O B E S     E - N E W S

Vol. 2, No. 6          July 6, 2001


This monthly newsletter is keep you informed about techniques to improve your immunogold labeling, highlight interesting articles and novel metal nanoparticle applications, and answer your questions. We hope you enjoy it and find it useful.

Have questions, or issues you would like to see addressed in the next issue? Let us know by e-mailing [email protected].

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Gold Lipids: Localize and track your liposomal formulations

Nanogold-labeled lipids are a unique probe from Nanoprobes, containing the 1.4 nm Nanogold cluster linked to palmitoyl or dipalmitoyl phosphatidyl ethanolamine moieties. By incorporating these compounds into liposomal preparations, you can make gold-labeled liposomes, and use them to localize and track drugs or other encapsulated agents in biological systems. For an example showing how this method was used to follow the delivery of an anti-fungal drug, see Adler-Moore, Bone Marrow Transplantation, 14, S3-S7 (1994)).

Nanogold labeling has also been used to demonstrate the targeting of cationic liposomes to endothelial cells in tumors and chronic inflammation; see Thurston, G., McLean, J. W., Rizen, M., Baluk, P., Haskell, A., Murphy, T. J., Hanahan, D., and McDonald, D. M.: J. Clin. Invest., 101, 1401-1413 (1998).

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Combined Fluorescent and Gold Probes: Check Labeling before EM Prep, or Obtain Correlative Labeling

FluoroNanogold is a unique probe containing both Nanogold® and fluorescein. You can use it for truly correlative labeling on the cellular and macromolecular level, or for checking your labeling by fluorescence before EM processing.

Catalog information: www.nanoprobes.com/FluoroN.html

Because FluoroNanogold combines two labels which are frequently used in different concentrations under different conditions, some optimization may be required to obtain the best performance in your experiments. For tips and tricks on FluoroNanogold labeling, and how to obtain the cleanest signal, visit our technical help page:

Technical help for FluoroNanogold: www.nanoprobes.com/TechFN.html

Original papers:

Powell, R. D.; Halsey, C. M. R.; Spector, D. L.; Kaurin, S. L.; McCann, J.;, and Hainfeld, J. F. A covalent fluorescent-gold immunoprobe: "simultaneous" detection of a pre-mRNA splicing factor by light and electron microscopy. J. Histochem. Cytochem., 45, 947-956 (1997).
Reprint: http://www.jhc.org/cgi/reprint/45/7/947

Robinson, J. M., and Vandr, D. D. Efficient immunocytochemical labeling of leukocyte microtubules with FluoroNanogold: An important tool for correlative microscopy. J. Histochem. Cytochem., 45, 631-642 (1997).
Reprint: http://www.jhc.org/cgi/reprint/45/5/631

Demonstration of correlative labeling:

Takizawa, T.; Suzuki, K., and Robinson, J. M.: Correlative Microscopy Using FluoroNanogold on Ultrathin Cryosections: Proof of Principle; J. Histochem. Cytochem., 46, 1097-1102 (1998).
Reprint: http://www.jhc.org/cgi/reprint/46/10/1097.pdf

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Conjugate gold particles to DNA

You can conjugate gold particles to oligonucleotides using several approaches. If you can incorporate a unique reactive group, such as a thiol, during oligonucleotide synthesis, you can label the thiolated oligonucleotide with Nanogold labeling reagents. See:

Alivisatos, A. P.; Johnsson, K. P.; Peng, X.; Wilson, T. E.; Loweth, C. J.; Bruchez, M. P., Jr., and Schultz, P. G.: Nature, 382, p. 609 (1996).
Abstract: http://www.ncbi.nlm.nih.gov:80/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=8757130&dopt=Abstract

Dubertret and co-workers used the same approach to label molecular beacons; see

Dubertret, B., Calame, M., and Libchaber, A. Nat. Biotechnol., 19, 365-370 (2001).
Abstract: http://www.ncbi.nlm.nih.gov:80/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=11283596&dopt=Abstract

Alternative strategies are available for DNA labeling: www.nanoprobes.com/TechNGlr.html#DNA

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Technical Help

If you have questions about gold labeling or silver enhancement, or need a solution for a problem, there's a good chance you can find the answer in our technical help pages. We have advice for obtaining the highest labeling, how to get the reagents to dissolve better, and how to optimize your gold labeling and silver enhancement to remove background problems. For more detailed information about how Nanogold and undecagold labeling works, check out our Guide to Gold Cluster Labeling. For references and descriptions of protocols that our users have developed with Nanogold, check our review articles in the Journal of Histochemistry and Cytochemistry (Hainfeld, J. F., and Powell, R. D.: J. Histochem. Cytochem., 48, 471-480 (2000)), or Cell Vision (Hainfeld, J. F., and Powell, R. D.: Cell Vision, 4, 408-432 (1997)).

E-mail us ([email protected]) if you would like a copy of the Cell Vision paper.

Abstract - J. Histochem. Cytochem.: http://www.jhc.org/cgi/content/abstract/48/4/471

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Recent Papers

The final version of the paper by Stewart and co-workers, describing the use of Nanogold to localize the two dimers of L7/L12 within the structure of the 70S ribosome, is published. Protein L7/L12 was reduced with 1 % mercaptoethanol and labeled with Monomaleimido Nanogold; two reconstitution approaches, together with cryo-electron microscopy and single particle reconstruction, were used to determine the structure.

Reference:

Montesano-Roditis, L.; Glitz, D. G.; Traut, R. R., and Stewart, P. L.: Cryo-electron Microscopic Localization of Protein L7/L12 within the Escherichia coli 70 S Ribosome by Difference Mapping and Nanogold Labeling. J. Biol. Chem., Vol. 276, Issue 17, 14117-14123, April 27, 2001.

If you are interested in labeling RNA with Nanogold, Medalia and co-workers have described a useful approach. They used ribonucleoside triphosphate analogs with a terminal thiol group attached to the heterocyclic ring for the in vitro transcription of RNAs carrying free thiol groups; these were then labeled with Monomaleimido- Nanogold. The complete procedure is given in their paper:

Medalia, O.; Heim, M.; Guckenberger, R.; Sperling, R., and Sperling, J.: Gold-Tagged RNA - A Probe for Macromolecular Assemblies. J. Struct. Biol., 127, 113-119 (1999).

Abstract: http://www.idealibrary.com/links/doi/10.1006/jsbi.1999.4134

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