Purpose: This study aimed to evaluate the effectiveness of low-fidelity simulation and actual human practice on the learning outcomes of a peripheral intravenous injection program for nursing students. Methods: Seventy-two third-year nursing students from a university in Seoul were included and divided into two groups; 38 and 34 in the low-fidelity simulation and human practice groups, respectively. The program was conducted from June 19–20, 2024. Data were analyzed using frequency, percentage, mean, standard deviation, χ²-test, independent t-test, and ANCOVA. Results: Both groups showed significant improvements in venipuncture performance, communication skills, empathy, academic performance, and self-efficacy. However, there were no statistically significant differences in these outcomes between both groups. Additionally, post-program learning satisfaction was similarly high in both groups. Conclusion: Low-fidelity simulation is as effective as actual human practice in achieving comparable learning outcomes in a peripheral intravenous injection program for nursing students. With appropriate instructional design, low-fidelity simulation can be a viable alternative to human practice, providing flexibility and accessibility in nursing education. This study underscores the importance of incorporating additional educational modules that enhance emotional and social skills, such as communication and empathy, tailored to the characteristics of individual learners.