A Reproducible Protocol for Embryonic Chicken Dorsal Root Ganglion Explant Culture and Quantitative Neurite Outgrowth Analysis

Background: Primary embryonic dorsal root ganglion (DRG) explants offer an accessible and physiologically relevant organotypic in vitro assay platform that preserves native three-dimensional cellular interactions and supports robust neurite outgrowth, enabling studies of sensory neuron development and neurite outgrowth, as well as controlled assessment of neurotoxicity, and pharmacological compound effects. Methods: Here we provide a comprehensive step-by-step method to micro-dissect and culture embryonic chicken DRGs and quantitatively measure neurite outgrowth. The protocol covers embryo preparation, vertebral column exposure, bilateral DRG microdissection, explant culturing, longitudinal phase-contrast imaging, and neurite image analysis. Results: DRG explants maintained their overall ganglion morphology and remained attached to the culture surface throughout the five-day culture period. Untreated control explants exhibited limited neurite outgrowth, whereas nerve growth factor (NGF) treated explants developed progressively longer and denser radial neurite networks. Neurite outgrowth was readily visualized by live phase contrast microscopy and quantified using an Image-based neurite tracing method, demonstrating the responsiveness of the explant model to NGF and its suitability for quantitative assessment of neurite outgrowth. Conclusion: This simple, low-cost protocol provides an in vitro assay platform for neurotoxicity, neuroregeneration, drug screening, and biomaterial evaluation.