Functional quantification of cyclosporine A and FK506 in human whole blood by flow cytometry, using the green fluorescent protein as an interleukin-2 reporter gene

2001 ◽  
Vol 256 (1-2) ◽  
pp. 77-87 ◽  
Author(s):  
Jean-Luc Taupin ◽  
Pierre Merville ◽  
Tarun McBride ◽  
Luc Potaux ◽  
Jean-François Moreau
Keyword(s):  
Flow Cytometry ◽  
Green Fluorescent Protein ◽  
Reporter Gene ◽  
Cyclosporine A ◽  
Whole Blood ◽  
Fluorescent Protein ◽  
Interleukin 2 ◽  
Human Whole Blood ◽  
Green Fluorescent

In vitro and in vivo evaluation of a green fluorescent protein-HSV1-TK dual-function pet reporter gene

2001 ◽  
Vol 44 (S1) ◽  
pp. S339-S341
Author(s):  
K. E. Luker ◽  
G. D. Luker ◽  
C. M. Pica ◽  
J. L. Dahlheimer ◽  
T. J. Fahrner ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Reporter Gene ◽  
Fluorescent Protein ◽  
Dual Function ◽  
In Vivo Evaluation ◽  
Green Fluorescent

GATA-1 expression pattern can be recapitulated in living transgenic zebrafish using GFP reporter gene

Development ◽  
1997 ◽  
Vol 124 (20) ◽  
pp. 4105-4111 ◽  
Author(s):  
Q. Long ◽  
A. Meng ◽  
H. Wang ◽  
J.R. Jessen ◽  
M.J. Farrell ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Progenitor Cells ◽  
Reporter Gene ◽  
Fluorescent Protein ◽  
Expression Patterns ◽  
Transgenic Fish ◽  
Green Fluorescent Protein Reporter ◽  
Erythroid Progenitor ◽  
Green Fluorescent ◽  
Fluorescent Protein Reporter

In this study, DNA constructs containing the putative zebrafish promoter sequences of GATA-1, an erythroid-specific transcription factor, and the green fluorescent protein reporter gene, were microinjected into single-cell zebrafish embryos. Erythroid-specific activity of the GATA-1 promoter was observed in living embryos during early development. Fluorescent circulating blood cells were detected in microinjected embryos 24 hours after fertilization and were still present in 2-month-old fish. Germline transgenic fish obtained from the injected founders continued to express green fluorescent protein in erythroid cells in the F1 and F2 generations. The green fluorescent protein expression patterns in transgenic fish were consistent with the pattern of GATA-1 mRNA expression detected by RNA in situ hybridization. These transgenic fish have allowed us to isolate, by fluorescence-activated cell sorting, the earliest erythroid progenitor cells from developing embryos for in vitro studies. By generating transgenic fish using constructs containing other zebrafish promoters and green fluorescent protein reporter gene, it should be possible to visualize the origin and migration of any lineage-specific progenitor cells in a living embryo.

Comparative production of human interleukin-2 fused with green fluorescent protein in several recombinant expression systems

2005 ◽  
Vol 24 (3) ◽  
pp. 225-233 ◽  
Author(s):  
Hyung Joon Cha ◽  
Hwa Sung Shin ◽  
Hye Jung Lim ◽  
Hye Sook Cho ◽  
Nimish N. Dalal ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Recombinant Expression ◽  
Interleukin 2 ◽  
Expression Systems ◽  
Green Fluorescent

scholarly journals Development of a Keratinocyte-Based Screening Model for Antipsoriatic Drugs Using Green Fluorescent Protein Under the Control of an Endogenous Promoter

2002 ◽  
Vol 7 (4) ◽  
pp. 325-332 ◽  
Author(s):  
Arno Pol ◽  
Fred Van Ruissen ◽  
Joost Schalkwijk
Keyword(s):  
Green Fluorescent Protein ◽  
Fusion Protein ◽  
Cell Line ◽  
Reporter Gene ◽  
High Throughput ◽  
Fluorescent Protein ◽  
Endogenous Promoter ◽  
Keratinocyte Cell ◽  
Egfp Fusion Protein ◽  
Green Fluorescent

Inflamed epidermis (psoriasis, wound healing, ultraviolet-irradiated skin) harbors keratinocytes that are hyperproliferative and display an abnormal differentiation program. A distinct feature of this so-called regenerative maturation pathway is the expression of proteins such as the cytokeratins CK6, CK16, and CK17 and the antiinflammatory protein SKALP/elafin. These proteins are absent in normal skin but highly induced in lesional psoriatic skin. Expression of these genes can be used as a surrogate marker for psoriasis in drug-screening procedures of large compound libraries. The aim of this study was to develop a keratinocyte cell line that contained a reporter gene under the control of a psoriasis-associated endogenous promoter and demonstrate its use in an assay suitable for screening. We generated a stably transfected keratinocyte cell line that expresses enhanced green fluorescent protein (EGFP), under the control of a 0.8-kb fragment derived from the promoter of the SKALP/elafin gene, which confers high levels of tissue-specific expression at the mRNA level. Induction of the SKALP promoter by tumor necrosis factor-ca resulted in increased expression levels of the secreted SKALP-EGFP fusion protein as assessed by direct readout of fluorescence and fluorescence polarization in 96-well cell culture plates. The fold stimulation of the reporter gene was comparable to that of the endogenous SKALP gene as assessed by enzyme-linked immunosorbent assay. Although the dynamic range of the screening system is limited, the small standard deviation yields a Z factor of 0.49. This indicates that the assay is suitable as a high-throughput screen, and provides proof of the concept that a secreted EGFP fusion protein under the control of a physiologically relevant endogenous promoter can be used as a fluorescence-based high-throughput screen for differentiation-modifying or antiinflammatory compounds that act via the keratinocyte.

scholarly journals Retrovirus-mediated in vitro gene transfer into chicken male germ line cells

Reproduction ◽  
2007 ◽  
Vol 134 (3) ◽  
pp. 445-453 ◽  
Author(s):  
Jiří Kalina ◽  
Filip Šenigl ◽  
Alena Mičáková ◽  
Jitka Mucksová ◽  
Jana Blažková ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Reporter Gene ◽  
Fluorescent Protein ◽  
Germ Line ◽  
Retroviral Vector ◽  
Male Germ Line ◽  
Testicular Cells ◽  
Infected Cells ◽  
Green Fluorescent ◽  
Transgenic Chickens

Chicken testicular cells, including spermatogonia, transplanted into the testes of recipient cockerels sterilized by repeated γ-irradiation repopulate the seminiferous epithelium and resume the exogenous spermatogenesis. This procedure could be used to introduce genetic modifications into the male germ line and generate transgenic chickens. In this study, we present a successful retroviral infection of chicken testicular cells and consequent transduction of the retroviral vector into the sperm of recipient cockerels. A vesicular stomatitis virus glycoprotein G-pseudotyped recombinant retroviral vector, carrying the enhanced green fluorescent protein reporter gene was applied to the short-term culture of dispersed testicular cells. The efficiency of infection and the viability of infected cells were analyzed by flow cytometry. No significant CpG methylation was detected in the infected testicular cells, suggesting that epigenetic silencing events do not play a role at this stage of germ line development. After transplantation into sterilized recipient cockerels, these retrovirus-infected testicular cells restored exogenous spermatogenesis within 9 weeks with approximately the same efficiency as non-infected cells. Transduction of the reporter gene encoding the green fluorescent protein was detected in the sperms of recipient cockerels with restored spermatogenesis. Our data demonstrate that, similarly as in mouse and rat, the transplantation of retrovirus-infected spermatogonia provides an efficient system to introduce genes into the chicken male germ line.

scholarly journals Constitutive and Inducible Expression of Green Fluorescent Protein in Brucella suis

1999 ◽  
Vol 67 (12) ◽  
pp. 6695-6697 ◽  
Author(s):  
Stephan Köhler ◽  
Safia Ouahrani-Bettache ◽  
Marion Layssac ◽  
Jacques Teyssier ◽  
Jean-Pierre Liautard
Keyword(s):  
Flow Cytometry ◽  
Green Fluorescent Protein ◽  
Fluorescence Microscopy ◽  
Gene Fusion ◽  
Fluorescent Protein ◽  
Fusion System ◽  
Chromosomal Dna ◽  
Brucella Suis ◽  
Green Fluorescent ◽  
Inducible Genes

ABSTRACT A gene fusion system based on plasmid pBBR1MCS and the expression of green fluorescent protein was developed for Brucella suis, allowing isolation of constitutive and inducible genes. Bacteria containing promoter fusions of chromosomal DNA togfp were visualized by fluorescence microscopy and examined by flow cytometry. Twelve clones containing gene fragments induced inside J774 murine macrophages were isolated and further characterized.

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