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Abstract Tertiary lymphoid structures (TLSs) provide specialized niches for immune cells, resulting in improved prognoses for patients undergoing cancer immunotherapy. Shaping TLS‐like niches may improve anti‐cancer immunity and overcome the current limitations of immune cell‐based immunotherapy. Here, it is shown that stromal vascular fraction (SVF) from adipose tissues can enhance dendritic cell (DC)‐mediated T cell immunity by inducing ectopic T lymphocyte clusters. SVF cells expanded ex vivo have phenotypes and functions similar to those of fibroblastic reticular cells in a secondary lymphoid organ, and their properties can be modulated using three‐dimensional spheroid culture and coculture with DCs spiked with antigen‐loaded iron oxide–zinc oxide core‐shell nanoparticles. Thereby, the combination of SVF spheroids and mature DCs significantly augments T cell recruitment and retention at the injection site. This strategy elicits enhanced antigen‐specific immune response and anti‐tumoral immunity in mice, illustrating the potential for a novel immunotherapeutic design using SVF as a structural scaffold for TLS.