Cellular GFP Toxicity and Immunogenicity: Potential Confounders in in Vivo Cell Tracking Experiments

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Abstract

Green Fluorescent protein (GFP), used as a cellular tag, provides researchers with a valuable method of measuring gene expression and cell tracking. However, there is evidence to suggest that the immunogenicity and cytotoxicity of GFP potentially confounds the interpretation of in vivo experimental data. Studies have shown that GFP expression can deteriorate over time as GFP tagged cells are prone to death. Therefore, the cells that were originally marked with GFP do not survive and cannot be accurately traced over time. This review will present current evidence for the immunogenicity and cytotoxicity of GFP in in vivo studies by characterizing these responses.

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Most cited references 49

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Improving the photostability of bright monomeric orange and red fluorescent proteins.

Nathan Shaner, Michael Lin, Michael R. McKeown … (2008)

All organic fluorophores undergo irreversible photobleaching during prolonged illumination. Although fluorescent proteins typically bleach at a substantially slower rate than many small-molecule dyes, in many cases the lack of sufficient photostability remains an important limiting factor for experiments requiring large numbers of images of single cells. Screening methods focusing solely on brightness or wavelength are highly effective in optimizing both properties, but the absence of selective pressure for photostability in such screens leads to unpredictable photobleaching behavior in the resulting fluorescent proteins. Here we describe an assay for screening libraries of fluorescent proteins for enhanced photostability. With this assay, we developed highly photostable variants of mOrange (a wavelength-shifted monomeric derivative of DsRed from Discosoma sp.) and TagRFP (a monomeric derivative of eqFP578 from Entacmaea quadricolor) that maintain most of the beneficial qualities of the original proteins and perform as reliably as Aequorea victoria GFP derivatives in fusion constructs.

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Fusion of bone-marrow-derived cells with Purkinje neurons, cardiomyocytes and hepatocytes.

Manuel Alvarez-Dolado, Ricardo Pardal, Jose M Garcia-Verdugo … (2003)

Recent studies have suggested that bone marrow cells possess a broad differentiation potential, being able to form new liver cells, cardiomyocytes and neurons. Several groups have attributed this apparent plasticity to 'transdifferentiation'. Others, however, have suggested that cell fusion could explain these results. Using a simple method based on Cre/lox recombination to detect cell fusion events, we demonstrate that bone-marrow-derived cells (BMDCs) fuse spontaneously with neural progenitors in vitro. Furthermore, bone marrow transplantation demonstrates that BMDCs fuse in vivo with hepatocytes in liver, Purkinje neurons in the brain and cardiac muscle in the heart, resulting in the formation of multinucleated cells. No evidence of transdifferentiation without fusion was observed in these tissues. These observations provide the first in vivo evidence for cell fusion of BMDCs with neurons and cardiomyocytes, raising the possibility that cell fusion may contribute to the development or maintenance of these key cell types.

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Extraction, Purification and Properties of Aequorin, a Bioluminescent Protein from the Luminous Hydromedusan,Aequorea

Osamu Shimomura, Frank H. Johnson, Yo Saiga (1962)

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PubMed ID:: 27435468

PMC ID:: 5050239

DOI:: 10.1007/s12015-016-9670-8

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This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.

ScienceOpen disciplines: Molecular medicine

Keywords: green fluorescent protein (gfp),cytotoxicity,immunogenicity,reporter gene,in vivo cell tracking,cell death

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ScienceOpen disciplines: Molecular medicine

Keywords: green fluorescent protein (gfp), cytotoxicity, immunogenicity, reporter gene, in vivo cell tracking, cell death

Cellular GFP Toxicity and Immunogenicity: Potential Confounders in in Vivo Cell Tracking Experiments

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Abstract

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Neuronal Signaling

Most cited references 49

Improving the photostability of bright monomeric orange and red fluorescent proteins.

Fusion of bone-marrow-derived cells with Purkinje neurons, cardiomyocytes and hepatocytes.

Extraction, Purification and Properties of Aequorin, a Bioluminescent Protein from the Luminous Hydromedusan,Aequorea

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