New two-layer Ruddlesden—Popper (RP) oxide La_0.25Sr_2.75FeNiO_7−δ (LSFN) in the combination of Sr₃Fe₂O_7−δ and La₃Ni₂O_7−δ was successfully synthesized and studied as the potential active single-phase and composite cathode for protonic ceramics fuel cells (PCFCs). LSFN with the tetragonal symmetrical structure (I4/mmm) is confirmed, and the co-existence of Fe³⁺/Fe⁴⁺ and Ni³⁺/Ni²⁺ couples is demonstrated by X-ray photoelectron spectrometer (XPS) analysis. The LSFN conductivity is apparently enhanced after Ni doping in Fe-site, and nearly three times those of Sr₃Fe₂O_7−δ, which is directly related to the carrier concentration and conductor mechanism. Importantly, anode supported PCFCs using LSFN-BaZr_0.1Ce_0.7Y_0.2O_3−δ (LSFN-BZCY) composite cathode achieved high power density (426 mW·cm⁻² at 650 °C) and low electrode interface polarization resistance (0.26 Ω·cm²). Besides, distribution of relaxation time (DRT) function technology was further used to analyse the electrode polarization processes. The observed three peaks (P1, P2, and P3) separated by DRT shifted to the high frequency region with the decreasing temperature, suggesting that the charge transfer at the electrode-electrolyte interfaces becomes more difficult at reduced temperatures. Preliminary results demonstrate that new two-layer RP phase LSFN can be a promising cathode candidate for PCFCs.