Objective: Clinical applications of human stem cells require large-scale expansion under chemically defined conditions. Appropriate in vitro culture systems should include synthetic substrates. Large-scale applications of expensive materials, such as peptide-based substrates, are cost-prohibitive. Materials and Methods: Coatings consisting of polymerized 3,4-dihydroxy-L-phenylalanine (DOPA) were applied to flat synthetic membranes and synthetic microcarriers. Large-scale expansion of undifferentiated human embryonic stem cells (hESC), induced pluripotent stem cells (hiPSC) and mesenchymal stem cells (hMSC) on these substrates was addressed by using serum- and xeno-free media. Results: The results revealed that the different types of human stem cells used could be propagated for up to at least 10 passages in the undifferentiated state on DOPAcoated susbtrates. Stem cell performance was comparable on commercial synthetic stem cell substrates and DOPA-coated substrates. DOPA-coated substrates are highly cost-effective in comparison to other synthetic substrates or Matrigel. Conclusion: We have developed cost-effective, fully synthetic substrates for the large-scale expansion of various types of human stem cells under conditions that are free from components purified from humans or animals.