Corrugated metallic surfaces offer means for efficient amplification of fluorescence bioassay signal based on the near field coupling between surface plasmons and fluorophore emitters that are used as labels. This paper discusses the design of such plasmonic structure to enhance the sensitivity of immunoassays with epi-fluorescence readout geometry. In particular, crossed gold grating is theoretically and experimentally investigated for combined increasing of the excitation rate at the fluorophore excitation wavelength and utilizing directional surface plasmon-coupled fluorescence emission. For Alexa Fluor 647 dye, the enhancement factor of around EF = 102 was simulated and experimentally measured. When applied to a sandwich interleukin-6 immunoassay, highly surface-selective enhancement reaching a similar value was observed. Besides increasing the measured fluorescence signal associated with the molecular binding events on a surface by two orders of magnitude, the presented approach enables measuring kinetics of the surface reaction that is otherwise masked by strong background signal originating from bulk solution.