The IncuCyte® NeuroBurst Orange is a lentiviral based live-cell neuronal labeling reagent driven by a synapsin promoter, resulting in the long term expression of a genetically encoded orange fluorescent calcium indicator (mRuby-based) in neuronal cells. The NeuroBurst Orange Reagent ensures highly efficient, yet non-disruptive labeling of primary or iPSC-derived neurons over days and weeks, and enables the kinetic quantification of neuronal activity and functional connectivity in the presence or absence of astrocytes. The NeuroBurst Orange Reagent has been validated for use with the IncuCyte® S3 Live-Cell Analysis System for Neuroscience (Cat. No. 4763) for measurements of neuronal activity and functional connectivity with the IncuCyte® S3 Neuronal Activity Analysis Software Module (Cat. No. 9600-0032). The IncuCyte S3 for Neuroscience is configured with an Orange/NIR Optical Module. The NeuroBurst Orange Reagent is NOT compatible with IncuCyte instruments configured with a Green/Red Optical Module, e.g. Cat. No. 4647.
Efficient, non-perturbing labeling of living cells enables chronic, long-term neuronal activity analysis during network development. CNS.4U iPSC-derived neurons (Ncardia) seeded at 20,000 cells/well in a 96-well microplate were infected with NeuroBurst Orange Reagent to monitor neuronal activity over time. Movies (3 minute duration) captured on Day 15 and day 44 reveal spontaneous development of neuronal activity (calcium oscillations) while phase images enable qualitative inspection of cell morphology. Movies were taken for 3 minutes every 24 hours over the course of 45 days, reveal maintained expression of NeuroBurst Orange Reagent. Quantification of the orange fluorescent signal kinetics shows an increase in active object count over the course of the experiment along with a corresponding increase in mean correlation (connectivity) at later time points, signifying development of a functional, mature network.
Quick Guide: IncuCyte® NeuroBurst Orange Reagent