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Dual-Labeled Probes for Fluorescence and Electron Microscopy

Published online by Cambridge University Press:  02 July 2020

Richard D. Powell ,
Carol M. R. Halsey ,
Edmund Gutierrez ,
James F. Hainfeld  and
Frederic R. Furuya
Richard D. Powell
Affiliation:
Nanoprobes, Incorporated, Stony Brook, NY11790
Carol M. R. Halsey
Affiliation:
Nanoprobes, Incorporated, Stony Brook, NY11790
Edmund Gutierrez
Affiliation:
Nanoprobes, Incorporated, Stony Brook, NY11790
James F. Hainfeld
Affiliation:
Nanoprobes, Incorporated, Stony Brook, NY11790
Frederic R. Furuya
Affiliation:
Nanoprobes, Incorporated, Stony Brook, NY11790
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Extract

Fluorescein and the 1.4 nm Nanogold cluster have been separately attached to polyclonal antibody Fab’ fragments to generate combined fluorescent and gold secondary antibody probes (1, 2), and in comparisons with Nanogold, colloidal gold and fluorescein-labeled secondary antibodies, neither label was found to be significantly compromised by the presence of the other. The proximity to a gold label with which a fluorophore retains sufficient fluorescence emission intensity to be useful is limited by non-radiative fluorescence resonance energy transfer (FRET) (3). This is a function of the degree of overlap of the fluorophore emission spectrum with the metal particle absorbtion spectrum, and of the separation of the metal particle and the fluorophore (4); for an effective dual-labeled probe, separation should be greater than the Forster distance (3), at which 50 % of excited-state decay occurs by FRET.

Type
Recent Advances in Labeling Techniques
Information
Microscopy and Microanalysis , Volume 4 , Issue S2: Proceedings: Microscopy & Microanalysis '98, Microscopy Society of America 56th Annual Meeting, Microbeam Analysis Society 32nd Annual Meeting, Atlanta, Georgia July 12-16, 1998 , July 1998 , pp. 992 - 993
Copyright
Copyright © Microscopy Society of America

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References

References:

1.Powell, R. D. et al., J. Histochem. Cytochem. 45 (1997) 947.CrossRefGoogle Scholar
2.Robinson, J. M. and Vandré, D. D.. J. Histochem. Cytochem. 45 (1997) 631.CrossRefGoogle Scholar
3.Wu, P. and Brand, L.. Anal. Biochem. 218 (1994) 1.CrossRefGoogle Scholar
4.Forster., Th., Ann. Physik., 2 (1948) 55.CrossRefGoogle Scholar
5.Powell, R. D. et al., Proc. Ann MSA Meeting, 54 (1996) 892.Google Scholar
6.Tomalia, D. A., Naylor, A. M. and Goddard III, W. A., Angew. Chem. Int. Ed. Eng., 29 (1990), 138.CrossRefGoogle Scholar
7. This work was supported by NIH grants 2R44 GM49564 and 1R44 GM56090.Google Scholar