Although pre-exposure prophylaxis (PrEP) is a key tool in HIV prevention efforts, little is known about PrEP as a prevention strategy for criminal justice-involved (CJI) women. The purpose of this study was to examine multilevel factors shaping PrEP awareness and acceptability among CJI women. Between January 2017 and December 2017, we conducted 52 interviews with CJI women at high risk for HIV and stakeholders from the criminal justice (CJ) and public health (PH) systems. Interviews explored awareness of PrEP and the multilevel factors shaping PrEP acceptability. Data were analyzed using inductive thematic analysis and executive summaries. Atlas.ti facilitated analyses. The majority of CJI women (n = 27) were, on average, 41.3 years, from racial and ethnic minority groups (56% Black/African-American; 19% Latinx) and reported engaging in recent high-risk behavior (nearly 60% engaged in transactional sex, 22% reported ≥ 4 sexual partners, and 37% reported injection drug use). Of system stakeholders (n = 25), 52% represented the CJ sector. Although CJI women were generally unaware of PrEP, attitudes toward PrEP were enthusiastic. Barriers to PrEP acceptability included medication side effects (individual level); distrust in HIV prevention mechanisms (community level); lack of local HIV prevention efforts among high-risk women (public policy/HIV epidemic stage level). Factors promoting PrEP included perceived HIV risk (individual level); PrEP being an HIV prevention method that women can control without partner negotiation (social and sexual network level); and availability of public health insurance (community level). Despite low awareness of PrEP, CJI women expressed positive attitudes toward PrEP. To improve PrEP access for CJI women, implementation efforts should address barriers and leverage facilitators across multiple levels to be maximally effective.