Despite the important role that teachers’ professional competencies play, domain-specific models of competence as well as established instruments to measure such competencies are lacking (e.g., Blömeke, Zlatkin-Troitschanskaia, Kuhn, & Fege, 2013 ). For this reason, a domain-specific model of competence and an instrument to measure prospective business and economics teachers’ professional competence in the domain of accounting was developed. This article focuses on the measurement of professional knowledge, which is a key facet of teachers’ professional competence. A corresponding test instrument is introduced and its measurement quality is reported. The test instrument used at 24 German universities (N = 1.158) comprises 49 items, distributed among different booklets following a multi-matrix design. All items have well functioning parameter values. In accordance with our hypothesis, a two-dimensional model fits the data best. The reliabilities of .64 (content knowledge) and .64 (pedagogical content knowledge) are satisfying. Thus, the developed instrument allows to gain a detailed understanding of prospective teachers’ professional knowledge in accounting.