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  Rapid characterization of green fluorescent protein fusion proteins on the molecular and cellular level by fluorescence correlation microscopy

Brock, R., Vamosi, G., Vereb, G., & Jovin, T. M. (1999). Rapid characterization of green fluorescent protein fusion proteins on the molecular and cellular level by fluorescence correlation microscopy. Proceedings of the National Academy of Sciences USA, 96, 10123-10128.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0012-FA3E-C Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0027-E88E-A
Genre: Journal Article

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 Creators:
Brock, R.1, Author              
Vamosi, G., Author
Vereb, G.1, Author              
Jovin, T. M.2, Author
Affiliations:
1Department of Molecular Biology, MPI for biophysical chemistry, Max Planck Society, ou_578628              
2Max Planck Society, ou_persistent13              

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Free keywords: Animal; Automation; Cytoplasm/ul [Ultrastructure]; CHO Cells; Endoplasmic Reticulum/ul [Ultrastructure]; Hamsters; *Luminescent Proteins/an [Analysis]; Luminescent Proteins/ul [Ultrastructure]; Microscopy, Fluorescence/is [Instrumentation]; *Microscopy, Fluorescence/mt [Methods]; *Receptor, Epidermal Growth Factor/an [Analysis]; *Receptor, Epidermal Growth Factor/ul [Ultrastructure]; *Recombinant Fusion Proteins/an [Analysis]; *Recombinant Fusion Proteins/ul [Ultrastructure]; Sensitivity and Specificity; Transfection
 Abstract: Fluorescence correlation microscopy (FCM) was applied to characterize fusion proteins of the green fluorescent protein (GFP) on the cellular as well as molecular level within seconds in an integrated instrument. FCM combines the inherent sensitivity and high spatial resolution of fluorescence correlation spectroscopy with fluorescence imaging and micropositioning, thereby providing a spectrum of molecular information in the cellular context. Signatures of characteristic parameters derived from the autocorrelation functions served to distinguish a GFP fusion protein of the epidermal growth factor receptor from GFP fluorescence in the endoplasmic reticulum and cytoplasm. Diffusion constants measured for free transiently expressed GFP reproduced values reported previously with other techniques. The accessible concentration range extends from millions to only a few thousand molecules per cell, with single molecule detectability in the femtoliter detection volume. The detailed molecular characterization offered by FCM is fully compatible with automation in sample identification and detection, offering new possibilities for highly integrated high-throughput screening.

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Language(s): eng - English
 Dates: 2005-07-081999
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Method: -
 Identifiers: eDoc: 223395
Other: 11466
 Degree: -

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Title: Proceedings of the National Academy of Sciences USA
Source Genre: Journal
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Affiliations:
Publ. Info: -
Pages: - Volume / Issue: 96 Sequence Number: - Start / End Page: 10123 - 10128 Identifier: -