Nitrogen (N) and sulfur (S) co-doped carbonaceous material has shown remarkably improved performance in electrocatalysis, supercapacitor, and so on. While, the application of N, S co-doped carbon in electrochemical sensor was less investigated. Herein, N, S co-doped yolk-shell porous carbon microspheres (N, S-YSPCMSs) were synthesized by silica templating strategy and high-temperature pyrolysis method. The as-prepared N, S-YSPCMSs exhibited spherical morphologies, porous structures and high N, S-doping contents. Owing to the synergetic effect of the well-dispersed N,S-dopants, good conductivity of carbon skeleton, high surface area and pore volume of the microspheres, the N, S-YSPCMSs prepared at 900 degrees C showed good electrochemical performance for Pb(II) and with a linear range of 4-160 mu g L-1, the limit of detection is 0.49 mu g L-1 (S/N = 3). The electrochemical sensor was consequently applied in detecting Pb(II) in fish serum samples with satisfying recoveries, indicating the N, S-YSPCMSs-900 are promising electrode components for electroanalysis.