Background: Kidney transplantation is the therapeutic of choice for patients with kidney failure. While immunosuppressive drugs can control graft rejection, their use is associated with increased infections and cancer, and they do not effectively control chronic graft rejection. Cell therapy is an attractive strategy to minimize the use of pharmacological drugs. Methods: We recently developed a protocol to generate human monocyte-derived autologous tolerogenic dendritic cells (ATDCs) from healthy volunteers. Herein, we transferred the ATDC manufacturing protocol to a Good Manufacturing Practice (GMP)-compliant facility. Furthermore, we compared the phenotype and in vitro functions of ATDCs generated from patients with end-stage renal disease to those generated from healthy volunteers. Results: We describe the critical steps for GMP-compliant production of ATDCs and define the quality criteria required to allow release of the cell products. Furthermore, we showed that ATDCs generated from healthy volunteers and patients with kidney failure display the same tolerogenic profile based on their phenotype, resistance to maturation, and ability to modulate T-cell responses. Conclusions: Together, these results allowed us to define the production process and the quality criteria for the release of ATDCs before their administration in patients receiving a kidney transplant.