NanoLuc/h-coelenterazine: an efficient pair for bioluminescence imaging in vivo and in vivo

Introduction. Bioluminescent labeling of tumor cells is becoming the standard in preclinical studies of novel anti-cancer drugs. Despite the variety of luciferases, many are not suitable for in vivo imaging.

Aim: to compare different Nanolux substrates in vitro and demonstrate the feasibility of using the NLuc/h-coelenterazine pair for in vivo imaging of disseminated tumor cells using the IVIS Spectrum system.

Materials and methods. The target cell line and T-cells expressing chimeric antigen receptors (CAR) were obtained via lentiviral cell transduction. Luminescence was measured with a Luminoskan™ Microplate Luminometer, while in vivo imaging in an NSG mouse model of CAR T-cell therapy was performed using the IVIS Spectrum system.

Results. Various substrates were compared in vitro for luminescence brightness and stability.

Additionally, the use of h-coelenterazine in combination with Nluc luciferase was demonstrated for in vivo cell imaging in mice. A genetically modified Nalm6-NLuc-CopGFP cell line was successfully obtained.

Conclusion. The comparison of various Nluc substrates in vitro and in vivo identified an optimal enzyme/substrate pair, which can serve as a valuable tool for assessing efficacy, safety and toxicity of compounds and cellular products being developed for antitumor therapy. The plasmid encoding Nluc can also be used to modify other cell lines necessary for the development and characterization of new gene therapy approaches.

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