Objective: This study assessed undergraduate unhealthy substance use, levels of knowledge about opioid pharmacology, unhealthy use and safety, and students' perceptions about their level of knowledge. Participants: Participants were undergraduate students at a large, private university in the Northeast. Methods: This study employed a cross-sectional study design using an online survey. A convenience sample of undergraduate students (N = 317) described their actual and perceived levels of opioid knowledge and their self-reported substance use. Results: Objective answers to quiz-style questions about opioids showed knowledge deficits. Participants rated themselves as slightly less knowledgeable than the "average student." Ten percent of students surveyed had used opioids unhealthily at some point in their lifetime. Those reporting unhealthy opioid use had higher subjective and objective opioid knowledge scores. Conclusions: University students' subjective and objective knowledge of opioids warrants more effective and engaging substance use education on college campuses.