Amidst the COVID-19 pandemic, handwashing offers a simple and effective hygienic measure for disease prevention. Reportedly, a significant proportion of college students did not follow handwashing recommendations provided by the Centers for Disease Control and Prevention (CDC) in the pre-COVID era. The purpose of this cross-sectional study was to explore and explain the handwashing behavior among college students during the COVID-19 pandemic using a contemporary fourth-generation multi-theory model (MTM) of health behavior change. Data were collected from 713 college students at a large public university in the Southern U.S. in October 2020 using a validated 36-item survey. Statistical analyses included independent samples t-tests, Pearson correlation, and hierarchical regression modeling. Among students not following handwashing recommendations, the constructs of participatory dialogue (beta = 0.152; p < 0.05) and behavioral confidence (beta = 0.474; p < 0.0001) were statistically significant and accounted for 27.2% of the variance in the likelihood of initiation of the behavior. Additionally, the constructs of emotional transformation (beta = 0.330; p < 0.0001), practice for change (beta = 0.296; p < 0.0001), and changes in the social environment (beta = 0.180; p < 0.05) were statistically significant and accounted for 45.1% of the variance in the likelihood of sustaining handwashing behavior. This study highlights the applicability and usability of the MTM in designing and testing behavior change interventions and media messaging in campaigns targeting college students.