This article describes the development and validation of the Light and Spectroscopy Concept Inventory (LSCI), a 26-item diagnostic test designed (1) to measure students' conceptual understanding of topics related to light and spectroscopy, and (2) to evaluate the effectiveness of instructional interventions in promoting meaningful learning gains in an introductory college astronomy course. We also present the final field-tested version of the LSCI for general use by the astronomy education community.