Objectives/Goals: Men and women with opioid use disorder (OUD) show differences in their initiation, use patterns, and outcomes that may have biological underpinnings. Here we present data directly comparing adult male and female rats across heroin-induced behaviors in order to provide insight into the nuances of sex differences in OUD. Methods/Study Population: We first used a 6-hour intermittent access heroin self administration paradigm to quantify six distinct drug-taking and drug-seeking behaviors. Based on the sum of the z-scores for each behavior, we classified rats as having high- or low-severity phenotypes. In a separate group of rats, we adapted this classification system to a 10-minute continuous access self-administration paradigm to better represent the timeframe of use common in people. Finally, we examined locomotor sensitization following daily heroin injections in two groups of rats. The first were given 2 mg/kg/day i.p. heroin for 10 days and the second were given 0.55 mg/kg/day i.v. heroin for 20 days. Results/Anticipated Results: In the 6-hour intermittent access, both sexes showed variability across individuals, but a greater proportion of females were classified as having a high-severity phenotype compared to males. This difference in severity distributions was also found in the 1-hour continuous access experiment. Consistent with the literature, in our sensitization experiments, we found that males had a lower baseline level of locomotion compared to females. Across sex and route of administration, rats treated with heroin initially decreased locomotion, but returned to baseline over the course of treatment. Females given i.v. infusions showed a rapid escalation of locomotion past baseline that was not seen in males or following i.p. injections. Discussion/Significance of Impact: Our results indicate a consistent pattern of females having a greater behavioral response to heroin compared to males. This suggests a sex-based effect on OUD that may interact with gender-based influences. As such, future research needs to consider sex in the development of treatments for OUD and other substance use disorders.