Engineered gold nanostructured arrays with feature separation in the sub-10-nm range were fabricated and used for highly sensitive surface enhanced Raman scattering (SERS) detection of biomarkers. Nanostructuring is achieved by combining electron beam lithography with the so-called shadow evaporation technique which involves a two-step Au vapor deposition through a suspended Ge mask while the substrate is tilted in opposite directions. This results in a regular triangular surface pattern with extremely small gap distances that allow positive enhancement of the local electric fields by enabling improvements in the electromagnetic coupling between adjacent nanoparticles. The resulting SERS active surfaces are suitable for the realization of reference procedures for quantifying marker molecules like urea or creatinine at physiologically relevant concentrations.