Abstract: This project aims to develop a measurement tool for assessing the conceptual understanding of secondary grade-level biology students (ages 11 to 18) that is reliable and valid. The study reported here describes the validity assessment of Secondary Biology Concept Inventory (S-BCI). A pool of assessment tasks was designed to target major biology constructs. The assessment items’ answer stems were developed to include distractors representing students’ alternative conceptions obtained from literature and student interviews. The validation stage of the S-BCI development involved an iterative revision and review process to help establish sufficient S-BCI content and construct validity. This stage included (i) student interviews and (ii) multi-expert panel critique. Based on the results of the aforementioned analyses, assessment items were proven to be valid where included on the S-BCI.