Bioflocs are rich in a variety of probiotics and bioactive compounds, which have been documented to promote growth, regulate antioxidant status and enhance the immune system. A 60‐day experiment was conducted to investigate the effects on growth, antioxidant and immune status, NF‐κB/Nrf2 signalling molecules and stress resistance in juvenile Rhynchocypris lagowskii Dybowski fed different dietary biofloc supplementations. Four hundred fifty R. lagowskii were fed five experimental diets containing graded levels of biofloc from 0 to 160 g/kg (referred to as B0, B4, B8, B12 and B16, respectively). The results showed that biofloc supplementation markedly promoted growth performance (weight gain rate, specific growth rate); improved amylase, lipase and protease activity; and enhanced the activities of glutathione peroxidase, catalase, total superoxide dismutase, total antioxidant capacity, lysozyme, complement C3, complement C4 and immunoglobulin M in the gill, hepatopancreas, kidney, gut and serum. The malondialdehyde content was significantly reduced with treatment B12. In addition, the levels of heat shock protein 70, heat shock protein 90, interferon‐γ and insulin‐like growth factor I were substantially increased with treatment B12 compared to those with the control treatment. Furthermore, biofloc assisted in regulating the expression of NF‐κB/Nrf2 signalling molecule genes, including NF‐κB, TNF‐α, IL‐1β, IL‐8, IL‐10, TGF‐β, Nrf2, Keap1, Maf, HO‐1, CAT, GPX, GCLC and CuZn‐SOD. Overall, our results suggest that dietary supplementation with biofloc can promote growth, improve immune and antioxidant status, and enhance NF‐κB/Nrf2 signalling molecule expression and stress resistance in juvenile R. lagowskii. A suitable supplementation level of approximately 120 g/kg biofloc is recommended in the present study.