<p>Preschool teachers’ inclusive knowledge is imperative for addressing the diverse demands of children with special education needs and enhancing the quality of inclusive education. However, the measuring instruments of preschool teachers’ inclusive knowledge are relatively few and primarily rely on self-report scales and semi-structured interviews, highlighting the necessity to enhance the objectivity and accuracy of the measurement. The main objective of this study was to develop a reliable instrument for evaluating preschool teachers’ proficiency in inclusive education knowledge. This study included a total of 4784 samples from four different provinces and cities in China, utilizing the Rasch model to assess the psychometric properties of the questionnaire. The results suggested that the questionnaire adhered to the unidimensional structure assumed by the Rasch model, with all items demonstrating local independence. All 28 items exhibited satisfactory fit indices (infit MNSQ: 0.50 to 1.14, outfit MNSQ: 0.86 to 1.07), aligning closely with the expectation of the Rasch model. Furthermore, the difficulty parameters (β) of the items were relatively low, and the test contributed maximal information to participants with latent traits ranging from −2 to 0 (θ). The marginal reliability of the test was satisfactory, while the differential item functioning indicated that the test has acceptable generalizability validity. In general, the Preschool Teachers’ Inclusive Knowledge Questionnaire demonstrated robust psychometric properties and exhibited broad applicability among preschool teachers of diverse genders and employment statuses.</p>