Automated neuronal reconstruction with super-multicolour Tetbow labelling and threshold-based clustering of colour hues.

ABSTRACT: Fluorescence imaging is widely used for the mesoscopic mapping of neuronal connectivity. However, neurite reconstruction is challenging, especially when neurons are densely labelled. Here, we report a strategy for the fully automated reconstruction of densely labelled neuronal circuits. Firstly, we establish stochastic super-multicolour labelling with up to seven different fluorescent proteins using the Tetbow method. With this method, each neuron is labelled with a unique combination of fluorescent proteins, which are then imaged and separated by linear unmixing. We also establish an automated neurite reconstruction pipeline based on the quantitative analysis of multiple dyes (QDyeFinder), which identifies neurite fragments with similar colour combinations. To classify colour combinations

SUBMITTER: Leiwe MN 

PROVIDER: S-EPMC11199630 | biostudies-literature | 2024 Jun

REPOSITORIES: biostudies-literature