There are professional expectations for public health nurses to develop competencies in evidence-informed decision-making (EIDM) due to its potential for improved client outcomes. Robust tools to assess EIDM competence can encourage increased EIDM engagement and uptake. This study aimed to develop and validate the content of a measure to assess EIDM competence among public health nurses. A four-stage process, based on measure development principles and the Standards for Educational and Psychological Testing, was used to develop and refine items for a new EIDM competence measure: a) content coverage assessment of existing measures; b) identification of existing measures for use and development of items; c) validity assessment based on content; d) validity assessment based on response process. An EIDM competence measurement tool consisting of EIDM knowledge, skills, attitudes/beliefs, and behaviour items was developed using conceptual literature and existing measures (Evidence-Based Practice Competency Tool and Evidence-Based Practice Beliefs Scale) to address limitations of existing EIDM tools identified from the content coverage assessment. Item content validity index ratings ranged from 0.64–1.00. Qualitative themes from validity assessment based on content and response process included word changes to improve clarity, reducing item redundancy, separating multi-component items, and ensuring items reflect nursing role expectations. Upon determining its reliability and validity, there is potential for the EIDM competence measure to be used in: public health nursing practice to identify competence gaps and strengths to facilitate professional development activities; in research to support development of strategies to build EIDM capacity; and for curriculum planning and development across nursing education programs.