A covalent fluorescent-gold immunoprobe: simultaneous detection of a pre-mRNA splicing factor by light and electron microscopy

Powell, R. D., Halsey, C. M., Spector, D. L., Kaurin, S. L., McCann, J., Hainfeld, J. F. (1997) A covalent fluorescent-gold immunoprobe: simultaneous detection of a pre-mRNA splicing factor by light and electron microscopy. Journal of Histochemistry and Cytochemistry, 45 (7). pp. 947-956. ISSN 0022-1554

URL: http://www.ncbi.nlm.nih.gov/pubmed/9212820
DOI: 10.1177/002215549704500704

Abstract

Immunoprobes that combine a fluorescent label with a 1.4-nm gold cluster compound have been prepared by covalent conjugation with polyclonal antibody Fab' fragments. These new immunoconjugates allow the collection of two complementary sets of data, from fluorescence and electron microscopy, from a single labeling experiment. We find that incorporation of one or more fluorescein moieties into the coordinated ligands of the 1.4-nm Nanogold gold cluster label yields a stable, dual-function immunolabel in which fluorescence quenching is negligible. In a second synthetic strategy, Nanogold and fluorescein were separately covalently conjugated to Fab' fragments to yield a probe with very similar properties. With the use of Fab' fragments, the entire probe is smaller than a whole IgG molecule, and it exhibited excellent penetration properties. It was used to localize the pre-mRNA splicing factor SC35 in the HeLa cell nucleus by both fluorescence and electron microscopy.

Item Type: Paper
Uncontrolled Keywords: Fluorescein Fluoresceins Fluorescent Antibody Technique Fluorescent Dyes Gold HeLa Cells Humans Immunoglobulin Fab Fragments Microscopy, Immunoelectron Molecular Probes Nuclear Proteins RNA Splicing Ribonucleoproteins
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification
bioinformatics > genomics and proteomics > genetics & nucleic acid processing
bioinformatics > genomics and proteomics
Investigative techniques and equipment
Investigative techniques and equipment > microscopy > electron microscopy
Investigative techniques and equipment > microscopy
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > pre-mRNA
CSHL Authors:
Communities: CSHL labs > Spector lab
Depositing User: Matt Covey
Date Deposited: 10 Dec 2012 19:56
Last Modified: 29 Jan 2015 17:56
Related URLs:
URI: http://repository.cshl.edu/id/eprint/26339

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