Drawing on conservation of resources theory, this study aims to explore the mediating role of employees’ strengths use and the moderating role of employees’ ambitions in the relationship between empowering leadership and employees’ taking charge behaviors. A total of 333 Chinese employees from various organizations across China (including industries such as manufacturing, IT, and education) completed our survey at two time points, with a two-week lag. We used structural equation modeling and moderated mediation path analysis to test our hypotheses. The research revealed that strengths use partially mediated the relationship between empowering leadership and employees’ taking charge behaviors, and ambition positively moderated the relationship between empowering leadership and strengths use, as well as the indirect relationship between empowering leadership and employees’ taking charge behaviors through strengths use. It extends the application field of strengths use, providing a new theoretical perspective on behavioral mechanisms for understanding the empowering leadership–employees’ taking charge behaviors relationship, and offers valuable strategies for organizations and leaders to promote employees’ taking charge behaviors more effectively.