GFP-Nanobody-Based Fluorescence-Detection Size-Exclusion Chromatography (FSEC) Method for the Expression Screening of...

Membrane and secreted proteins are central to numerous biological processes and represent key therapeutic targets, yet their structural analysis remains hindered by challenges in expression and purification. Here, we present a streamlined screening strategy based on nanobody-assisted fluorescence detection that enables rapid assessment of protein quality directly from cell lysates or culture media. This approach overcomes limitations of traditional GFP-tagging methods and is broadly applicable to both membrane-spanning and secreted proteins. Using Low-density lipoprotein receptor-related protein 6 (Lrp6) as a case study, we demonstrate its utility for optimizing expression, guiding purification, and probing protein-protein interactions. This versatile platform provides a powerful tool for accelerating construct screening and advancing structural and functional studies.