Among biosensors of various types, the silicon nanowire field-effect transistor (SiNW-FET) is believed to be the most sensitive and powerful device for bio-applications. The principle of sensing is based on the variation of conductivity resulting from a disturbance of charge on the surface of the SiNW-FET, but this detection is feasible predominantly for charged analytes, such as a protein, DNA, antibody, virus etc. The objective of our work was to overcome this intrinsic weakness of a SiNW-FET and to develop a platform to detect steroids. For this purpose, we designed an engineered protein, Δ5-3-ketosteroid isomerase, to function as a steroid acceptor that was chemically modified with a carbon chain-linked 1,5-EDANS moiety, and further immobilized on the surface of a silicon nanowire. In the presence of a steroid, the negatively charged 1,5-EDANS moiety, which presumably occupies the steroid-binding site, is expelled and exposes to the nanowire surface. The electrical response produced from the 1,5-EDANS moiety is measured and the concentration is calculated accordingly. The sensitivity of this novel nano-bio-device can attain a femtomolar level.