Functional multiplex reporter assay using tagged Gaussia luciferase

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Abstract

We have developed a multiplex reporter system to monitor multiple biological variables in real-time. The secreted Gaussia luciferase was fused to ten different epitope tags (Gluc _tag), each expressed in different tumor cells. By immunobinding of the tags followed by Gluc _tag detection, this system allowed the independent and real-time monitoring of mixed cell cultures in vitro and of mixed subcutaneous and intracranial tumor subpopulations in vivo.

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Most cited references 7

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Intravital imaging.

Mikael J. Pittet, Ralph Weissleder (2011)

Until recently, the idea of observing life deep within the tissues of a living mouse, at a resolution sufficient to pick out cellular behaviors and molecular signals underlying them, remained a much-coveted dream. Now, a new era of intravital fluorescence microscopy has dawned. In this Primer, we review the technologies that made this revolution possible and demonstrate how intravital imaging is beginning to provide quantitative and dynamic insights into cell biology, immunology, tumor biology, and neurobiology. Copyright © 2011 Elsevier Inc. All rights reserved.

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Optical imaging of Renilla luciferase reporter gene expression in living mice.

S. Gambhir, S Bhaumik (2002)

Imaging reporter gene expression in living subjects is a rapidly evolving area of molecular imaging research. Studies have validated the use of reporter genes with positron emission tomography (PET), single photon emission computed tomography (SPECT), MRI, fluorescence with wild-type and mutants of green fluorescent protein, as well as bioluminescence using Firefly luciferase enzyme/protein (FL). In the current study, we validate for the first time the ability to image bioluminescence from Renilla luciferase enzyme/protein (RL) by injecting the substrate coelenterazine in living mice. A highly sensitive cooled charge-coupled device camera provides images within a few minutes of photon counting. Cells, transiently expressing the Rluc were imaged while located in the peritoneum, s.c. layer, as well as in the liver and lungs of living mice tail-vein injected with coelenterazine. Furthermore, d-luciferin (a substrate for FL) does not serve as a substrate for RL, and coelenterazine does not serve as a substrate for FL either in cell culture or in living mice. We also show that both Rluc and Fluc expression can be imaged in the same living mouse and that the kinetics of light production are distinct. The approaches validated will have direct applications to various studies where two molecular events need to be tracked, including cell trafficking of two cell populations, two gene therapy vectors, and indirect monitoring of two endogenous genes through the use of two reporter genes.

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Bioluminescence imaging: progress and applications.

Christian E Badr, Bakhos A Tannous (2011)

Application of bioluminescence imaging has increased tremendously in the past decade and has significantly contributed to core conceptual advances in biomedical research. This technology provides valuable means for monitoring of different biological processes in immunology, oncology, virology and neuroscience. In this review, we discuss current trends in bioluminescence and its application in different fields with an emphasis on cancer research. Copyright © 2011 Elsevier Ltd. All rights reserved.

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Author and article information

Journal

Journal ID (nlm-ta): Sci Rep

Journal ID (iso-abbrev): Sci Rep

Title: Scientific Reports

Publisher: Nature Publishing Group

ISSN (Electronic): 2045-2322

Publication date (Electronic): 10 January 2013

Publication date Collection: 2013

Volume: 3

Electronic Location Identifier: 1046

Affiliations

[1 ]Neuro-oncology Research Group, Department of Neurosurgery, Cancer Center Amsterdam, VU University Medical Center , Amsterdam, The Netherlands

[2 ]Department of Radiation Sciences, Oncology, Umeå University , Umeå, Sweden

[3 ]Neuroscience Center, Department of Neurology, Massachusetts General Hospital and Neuroscience Program, Harvard Medical School , Boston, MA, USA

Author notes

[a ] t.wurdinger@ 123456vumc.nl

[b ] Jonas.nilsson@ 123456onkologi.umu.se

Article

Publisher Item ID: srep01046

DOI: 10.1038/srep01046

PMC ID: 3541509

PubMed ID: 23308339

SO-VID: a224cbae-0614-4187-aba2-4cab76041ce9

License:

This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-nd/3.0/

Functional multiplex reporter assay using tagged Gaussia luciferase

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Functional role of amyloid

Most cited references 7

Intravital imaging.

Optical imaging of Renilla luciferase reporter gene expression in living mice.

Bioluminescence imaging: progress and applications.

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