A new surface-enhanced Raman scattering (SERS) active substrate has been developed based on our previous study. Small silver nanoparticles on a quartz slide can be enlarged by using a mixture of commercially available reagents called Silver Enhancer and Initiator. The optical properties and characteristics of the new substrate have been investigated by ultraviolet-visible (UV-Vis) spectroscopy and atomic force microscopy (AFM). The results indicate that the small silver nanoparticles grow and some silver aggregates emerge on the quartz slide after the slide is immersed into the Silver Enhancer and Initiator Mixture (SEIM). The average diameter of the silver nanoparticles on the substrate becomes approximately double after the immersion into SEIM for 20 s. 1,4-bis[2-(4-pyridyl)ethenyl]-benzene (BPENB) was used as a Raman probe to evaluate the enhancement ability of the new silver substrate. It has been found that the SERS intensity can be increased about 10 times by using the substrate treated by SEIM compared with that without being treated by SEIM. Interestingly enough, the SERS enhancement increases with time. This may be due to the reorganization of silver nanoparticles on the quartz surface. The new substrate can remain active for more than 90 days. The adsorption mode of BPENB on the new substrate and the dependence of the BPENB configurations on the concentration of BPENB in methanol solution have also been investigated by SERS or UV-Vis spectroscopy. The SERS spectra of a self-assembled monolayer (SAM) BPENB film adsorbed on a silver substrate treated by SEIM show that BPENB molecules are chemically absorbed through the Ag-N bond. Consequently, a nearly perpendicular orientation of BPENB on the silver surface is proposed. The SERS spectra of BPENB SAMs on the new substrates fabricated from methanol solutions with different concentrations are compared. The concentration dependence of the SERS spectra reveals that the BPENB molecules are adsorbed on the silver film as monomers when the film is prepared from the solution with a lower concentration (<4 x 10(-6) M) and as aggregates when it is prepared from the solution with a higher concentration (>1 x 10(-5) M).