This study conducts a comprehensive analysis of deer antler velvet's impact, with a specific emphasis on mouse models and in vitro experiments. The study navigates the intricacies of antler velvet's variability, encompassing considerations of drying methods, extraction techniques, and anatomical regions of antlers.Employing a diverse array of processing methods, this study prioritizes both food safety and the consistent intake of deer antler velvet extracts. The study scrupulously evaluates toxicity and immune-boosting properties through exhaustive assessments, utilizing Caenorhabditis elegans, immunosuppressive mouse models, and immune cells. The study unfolds the repercussions of deer antler velvet extract on the lifespan, neuromuscular functions, and cognitive abilities of C. elegans. Additionally, this study explores the extract's potential to alleviate toxicity induced by cyclophosphamide (CPA) in a mouse model, with a focus on inflammation, metabolic disorders, and gut microbiota composition. The antler velvet extract prevents immune dysregulation caused by CPA treatment by ameliorating systemic inflammation and restoring energy metabolism. Furthermore, antler velvet extract treatment significantly transforms the gut microbiota of an immunosuppressive mouse model by fostering the proliferation of commensal bacteria such as Lactobacillus, Akkermansia, and Lachnospiraceae at the genus level. Moreover, antler velvet treatment enhances the activity of natural killer cells against YAC-1 lymphoma while tempering overactivity among immune cells to baseline levels. In conclusion, this study provides nuanced insights into the diverse benefits of antler velvet extract, encouraging sustained research to unveil its complete potential in the realms of mouse models and in vitro experiments.