Substance abuse among working adults represents billions of dollars in preventable health care costs and industry financial loss. Therefore, it is imperative to develop and test effective substance abuse prevention programs for the workplace. However, applied workplace substance abuse prevention research is fraught with numerous methodological challenges. This article highlights a number of these challenges, which include (1) reaching a broad audience with prevention messages, (2) handling the concerns of the employer, (3) collecting substance use data in the workplace, (4) accessing and using records-based data, and (5) linking survey and records-based data. Using examples from the authors' ongoing research assessing a workplace health promotion and substance abuse prevention program, funded by the Substance Abuse and Mental Health Services Administration's Center for Substance Abuse Prevention, the authors address these challenges.