Fig. 2 shows the Raman spectra of R6G and MB Deltorphin II adsorbed on the SERS substrates prepared from the analyte solutions at 10–8 M. The characteristic Raman bands of R6G at 615, 775, 1127, 1182, 1310, 1362, 1508, 1573, and 1649 cm−1 (Fig. 2(1)  and ) and of MB at 495, 497, 677, 768, 890, 1067, 1181, 1331, 1396, 1411, 1544, and 1620 cm−1 were measured (Fig. 2(2) ). Raman spectra of R6G and MB prepared from the solutions with concentrations of 10−10 M and 10−9 M, were recorded over nine random spots to evaluate the uniformity in sensitivity of the SERS substrates, respectively. As displayed in Fig. 3, the R6G and MB Raman peaks were detected on all nine spots with similar peak intensities. The relative standard deviations (RSDs) were evaluated from the SERS spectra of the randomly selected nine spots for R6G and MB molecules and the RSD values for the main Raman peaks are summarized in Table 1. The averaged RSD values were determined to be about 15.93% and 5.86% for R6G and MB, respectively. The observation suggests that the analytes were uniformly distributed on the SERS substrates and the enhancement of the R6G and MB Raman signals with the substrates were highly reproducible over the SERS active area.