Evaluating reporter genes of different luciferases for optimized in vivo bioluminescence imaging of transplanted neural stem cells in the brain.

摘要: Bioluminescence imaging (BLI) has become the method of choice for optical tracking cells in small laboratory animals. However, use luciferases from different species, depending on substrates and emitting at distinct wavelengths, not been optimized sensitive neuroimaging. In order to identify most suitable luciferase, this quantitative study compared Luc2, CBG99, PpyRE9 hRluc. Human embryonic kidney (HEK-293) mouse neural stem were transduced by lentiviral vector-mediated transfer express one four luciferases, together with copGFP. A T2A peptide linker promoted stoichiometric expression between both reporters comparison cell populations upon flow cytometry. Cell dilution series used determine highest BLI sensitivity vitro Luc2. Coelenterazine h-dependent hRluc signals clearly exceeded d-luciferin-dependent vitro. For vivo analysis, transplanted into brain was performed including recording emission kinetics spectral characteristics. Differences light observed d-luciferin vs h. The spectra Luc2 remained almost unchanged, while spectrum CBG99 became biphasic. Most importantly, photon decreased feasibility combining dual color substrate neuroimaging tested discussed. This investigation provides first complete expressed cells. It results a clear recommendation as best luciferase selection