Reporter Gene Imaging by Purnima Dubey

Author:Purnima Dubey
Language: eng
Format: epub, pdf
Publisher: Springer New York, New York, NY

Jukka Pajarinen1, Tzu-Hua Lin1 and Stuart B. Goodman1

(1)Orthopaedic Research Laboratories, Department of Orthopaedic Surgery, Stanford University School of Medicine, Stanford, CA, USA

Stuart B. Goodman

Email: [email protected]

Abstract

Macrophages have emerged as crucial regulators of tissue homeostasis, inflammation, and tissue regeneration. In vivo bioluminescence imaging could offer a powerful tool to study many poorly understood aspects of macrophage biology. Thus, we recently developed a straightforward method for the production of large numbers of green fluorescent protein (GFP) and firefly luciferase (fLUC)-expressing reporter macrophages for various in vivo bioluminescence imaging applications. Lentivirus vector containing the GFP/fLUC reporter gene is produced and mouse bone marrow macrophages are isolated following established protocols. Macrophages are then exposed to the lentivirus in the presence of 10 μM cyclosporine for 24 h. After a 24-h recovery period, the transduction is repeated. Three days after the second infection the cells are ready to be used in vivo. Following this cyclosporine-mediated double infection strategy up to 60% of the macrophages express GFP in flow cytometry. The macrophages maintain their ability to polarize to M1 and M2 phenotypes and, when injected to the systemic circulation of a mouse model, reporter cells are both easily detectable with BLI and migrate to a local site of inflammation. These GFP/fLUC-expressing reporter macrophages could prove to be useful tools to study the role of macrophages in health and disease.

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