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A bio-based multi-layer building envelope assembly has been developed for its integration in newly built and retrofitted buildings. Forest-based materials and biocomposite profiles are used as an alternative to fossil-based insulants and metallic framing, providing a well-insulated and low-thermal-bridge technical solution. The wall assembly has been installed as the external envelope of one apartment of a housing block in Donostia-San Sebastián (Basque Country, Spain). A comparative study has been performed for the bio-based wall and the reference wall of the building. Their in-situ thermal resistance has been obtained by means of three different methods: (1) the steady-state average method, (2) a semi-dynamic method from heat balance at the internal surface, and (3) a dynamic multiple regression method. Reasonably consistent results have been obtained with the three methods: a discussion is provided on the influence of measuring periods and boundary conditions. Outputs from this experimental campaign are valuable as a counterpoint to desktop studies and tests under controlled laboratory conditions. Learnings and outputs from the present study should contribute to a better understanding of the in-situ performance of building envelope assemblies and their assessment methods.