Although surface-enhanced Raman spectroscopy (SERS) has recently emerged as a powerful analytical tool, its widespread use is hindered by several inherent problems (e.g., low reproducibility) that need to be solved before further progress is possible. Herein, we used dodecyltrimethylammonium bromide (DTAB), N-dodecyl-N-2-hydroxyethyl-N,N-dimethylammonium bromide (C 12 HDAB), and N-dodecyl-N,N-2-dihydroxyethyl-N-methylammonium bromide (C 12 DHAB) as templates for the seed-mediated growth of Au nanorods with different aspect ratios and probed the growth mechanism by a range of instrumental techniques. The prepared nanorods were used as substrates for SERS-based detection of three negatively charged colorants (lemon yellow, orange II, and amaranth) and exhibited extraordinary SERS sensitivity for all of these targets. The highest sensitivity was observed in the case of C 12 DHAB-capped nanorods, which was attributed to abundant hydrogen bonds between surfactants and targets and strong electrostatic adsorption of negatively charged colorants. Therefore, the above substrate was concluded to be well suited for real-life applications.