Critical thinking has been one of the 21st-century skills consistently associated with students' future career advancement as a positive student outcome of STEM education. The aim of the study is to develop and validate science critical thinking skill instruments to assess the improvement in the subject of living organisms and force and friction through design-based STEM education. In this design-based research study, the student's modules were developed by the integrated STEM education principles involving the activities and worksheets in line with the frame of critical thinking approach. The kappa statistics for content validity, exploratory and confirmatory factor analysis for construct validity, and item and reliability analysis for the quality of items were used in the development stage of instruments. The results of these analyses endorsed the 15 two-tier item for each test of Living Things Critical Thinking (LTCT) and Measuring Force and Friction Critical Thinking (MFFCT) as unidimensional constructs to produce valid and reliable data to measure the fifth grade students' critical thinking skills in the related science content. Comparing the pre and post applications of instruments in the study group indicated that STEM modules improved the students' science critical thinking skills such as interpretation, analysis, and inference. In this respect, developing and validating instruments to assess the integrated critical thinking skills will contribute to the empirical examination of this construct within the context of school science learning.