Background: Transitions in healthcare delivery, such as the rapidly growing numbers of older people and increasing social and healthcare needs, combined with nursing shortages has sparked renewed interest in differentiations in nursing staff and skill mix. Policy attempts to implement new competency frameworks and job profiles often fails for not serving existing nursing practices. This study is aimed to understand how licensed vocational nurses (VNs) and nurses with a Bachelor of Science degree (BNs) shape distinct nursing roles in daily practice. Methods: A qualitative study was conducted in four wards (neurology, oncology, pneumatology and surgery) of a Dutch teaching hospital. Various ethnographic methods were used: shadowing nurses in daily practice (65h), observations and participation in relevant meetings (n=56), informal conversations (up to 15 h), 22 semi-structured interviews and member-checking with four focus groups (19 nurses in total). Data was analyzed using thematic analysis. Results: Hospital nurses developed new role distinctions in a series of small-change experiments, based on action and appraisal. Our findings show that: (1) this developmental approach incorporated the nurses’ invisible work; (2) nurses’ roles evolved through the accumulation of small changes that included embedding the new routines in organizational structures; (3) the experimental approach supported the professionalization of nurses, enabling them to translate national legislation into hospital policies and supporting the nurses’ (bottom-up) evolution of practices. The new roles required the special knowledge and skills of Bachelor-trained nurses to support healthcare quality improvement and connect the patients’ needs to organizational capacity. Conclusions: Conducting small-change experiments, anchored by action and appraisal rather than by design, clarified the distinctions between vocational and Bachelor-trained nurses. The process stimulated personal leadership and boosted the responsibility nurses feel for their own development and the nursing profession in general. This study indicates that experimental nursing role development provides opportunities for nursing professionalization and gives nurses, managers and policymakers the opportunity of a ‘two-way-window’ in nursing role development, aligning policy initiatives with daily nursing practices.