The Karousis and Mühlemann labs developed based on dual centrifugation an efficient cell-free translation system for diverse human cells. They investigated several parameters in their protocol to assess their impact on the activity of the cell-free translation system. Their results have been published in the article “Efficient cell-free translation from diverse human cell types” in the Journal of Biological Chemistry.
Abstract
Cell-free translation systems are indispensable for studying protein synthesis, enabling researchers to explore translational regulation across different cell types. The difficulties in producing cell-free translation systems from different cell types limit the ability to study regulatory mechanisms that depend on different biological contexts. Here, we present a scalable method for preparing translation-competent lysates from a range of frequently used human cell lines using dual centrifugation. We optimized lysis conditions for adherent and suspension cells, producing high-quality lysates from HEK-293 (adherent and in suspension), HeLa, SH-SY5Y, and U2OS cells. Our results demonstrate that cell-specific factors influence translation efficiency, with adherent HeLa cells showing the highest activity. We also observed that sensitivity to lysis conditions varies between cell lines, underscoring the importance of fine-tuning parameters for efficient protein production. Our method provides a robust and adaptable approach for generating cell-type-specific lysates, broadening the application of in vitro translation systems in studying translational mechanisms.
Read the Publication in the Journal of Biological Chemistry (Open Access)
Abstract and figure from Ziegelmüller et al. (2025) J Biol Chem published under a CC BY 4.0 license.