Fraunhofer Institute for Translational Medicine and Pharmacology (ITMP) 1 article published in JoVE Bioengineering Fibroblast Derived Human Engineered Connective Tissue for Screening Applications Gabriela L. Santos1,2, Tim Meyer1,2, Malte Tiburcy1,2, Alisa DeGrave1,2, Wolfram-Hubertus Zimmermann1,2,3,4,5, Susanne Lutz1,2 1Institute of Pharmacology and Toxicology, University Medical Center Goettingen, 2DZHK (German Center for Cardiovascular Research) partner site, Goettingen, 3Cluster of Excellence “Multiscale Bioimaging: from Molecular Machines to Networks of Excitable Cells” (MBExC), University of Goettingen, 4Center for Neurodegenerative Diseases (DZNE), 5Fraunhofer Institute for Translational Medicine and Pharmacology (ITMP) Presented here is a protocol to generate engineered connective tissues for a parallel culture of 48 tissues in a multi-well plate with double poles, suitable for mechanistic studies, disease modeling, and screening applications. The protocol is compatible with fibroblasts from different organs and species and is exemplified here with human primary cardiac fibroblasts.