Olfactory sensory neurons (OSN) regenerate throughout life. Their renewal depends on the cellular architecture of the surrounding tissues and the molecular environment. As a consequence, in-vitro models to characterize OSN regeneration are often based on bulky olfactory epithelium (OE) biopsies or heterogenous cell cultures. To accurately quantify them, fluorescent OSN are usually manually traced to better discern each axon from other structures present in the culture. However, this manual procedure is time-consuming and prone to interpretation bias. To overcome this gap, we developed a specific and entirely automated neuronal growth quantification tool based on a MATLAB script operating on pictures of β-3 Tubulin stained OE biopsies. As a first step, the script enhances the contrast of the picture and detects the biopsy edges. These are used as a starting point to quantify axonal length. Only β-3 Tubulin positive pixels forming a tubular shape compatible with an axon radiating from the biopsy perimeter, are quantified using an algorithm called Fibermetric.The program reports the results (e.g. total axonal length, surface area of the biopsy) in an Excel file. Furthermore, it generates images of the processing steps and a scheme representing the probability of finding an axon at a given angle and distance relative to the biopsy. The comparison between automated and manual axonal quantification showed an intra class correlation coefficient (ICC) of 0.96 (p= 0.017). Furthermore, manual tracing took on average 23 minutes per each explant while the tool average computation time was 3 minutes. The script revealed itself significantly faster than the manual tracing (p= 0.0020). In conclusion, the new automated quantification tool provides an accurate, objective and time-saving method to measure OSN axonal outgrowth. It may facilitate high-throughput screens of regenerative compounds in the future. The project was founded by: Fondation Louis-Jeantet, Auris, Sir Jules Thorn.