One of the main challenges faced by a detection system is the ability to detect and identify trace amounts with high accuracy in real time. None of the methods used today is able to detect and identify trace levels of different sample types from small molecules to macro molecules, proteins, and large biological samples such as spores. Surface-enhanced Raman spectroscopy (SERS) is here demonstrated as a sensitive method for detection of chemical, biological, and explosive warfare agent simulants. Using a sealed chamber, vapor phase SER spectra of two C-simulants, 2-chloroethyl ethyl sulphide (CEES), dimethyl methylphosphonate (DMMP), and one test agent methylsalicylate (MES), can clearly be identified by comparison to their bulk Raman spectra in less than 40 seconds on a Klarite® nanostructured surface. Identification of low nanogram levels of TNT in 10 s on this commercially available nanostructured surface is a straight forward exercise. Two types of Bacillus spores, Bacillus thuringiensis and Bacillus atrophaeus, simulants for the anthrax spore, also gave enhanced Raman spectra on these surfaces, but better substrates are needed for reliable results that can be used for identification. Furthermore, freezing of colloidal gold nanoparticles is demonstrated as a method to produce aggregates without an aggregating agent and the achieved lower limit of detection of Rhodamine 6G is evaluated. Finally, a new type of nanostructured surface, gold and silver coated black silicon, is evaluated based on sensitivity for use in a portable SERS instrument under development of the company Serstech AB.