Contextually relevant factors within a training environment facilitate performance transfer to applied clinical settings. Live animals are often used for pediatric and neonatal intubation training because the scale of their airways and the living condition of the animal are assumed to facilitate transfer of aquired skills to intubating infants and children, despite species-specific anatomical variation. Mannequin simulators provide an alternative for training, but have anatomical, physiological, and tissue limitations. We evaluated the impact of two clinical training methods (live animal v. simulation) on the acquisition of performance abilities in pediatric and neonatal intubation. Methods: We implemented a quasi-experimental design with purposive sampling to assess performance differences between 294 subjects after completing a training intervention that included either direct interaction with a (1) live animal or (2) mannequin simulator. All other training elements were identical and followed American Heart Association (PALS/NRP) clinical protocols. We used validated instruments to assess knowledge, performance, and self-efficacy outcomes before and after training, and at three retention intervals (6, 18, and 52 weeks). Results: Post-training outcomes were significantly better that pre-training outcomes for both groups (p < .001), and there were no significant differences between the outcomes for the two types of training. There were significant differences between the performance outcomes after 18 and 52 weeks (p < .01), and for cognitive outcomes after 52 weeks (p < .01), all favoring the simulator training. Conclusions: These findings could aid in the selection of instructional methodologies that minimize the uses of live animals for instruction in pediatric and neonatal intubation without sacrificing the quality of training. Variation in retention outcomes are likely due to opportunity for repeated deliberate practice using a simulator rather than associated with fidelity issues. Improvements in simulator technology would likely improve training outcomes.