A very sensitive and highly specific heterogeneous immunoassay system, based on surface-enhanced Raman scattering (SERS) and gold nanoparticles, was developed for the detection of bacteria and other pathogens. Two different types of gold nanoparticles (citrate-stabilized gold nanosphere and hexadecyltrimethylammonium bromide (CTAB)-stabilized gold nanorod particles) were examined and this immunoassay was applied for the detection of Escherichia coli. Raman labels were constructed by using these spherical and rod-shaped gold nanoparticles which were first coated with 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) and subsequently with a molecular recognizer. The working curve was obtained by plotting the intensity of the SERS signal of the symmetric NO2 stretching of DTNB at 1,333 cm(-1) versus the concentration of the E. coli. The analytical performance of gold particles was evaluated via a sandwich immunoassay, and linear calibration graphs were obtained in the E. coli concentration range of 10(1)-10(5) cfu/mL with a 60-s accumulation time. The sensitivity of the Raman label fabricated with gold nanorods was more than three times higher than spherical gold nanoparticles. The selectivity of the developed sensor was examined with Enterobacter aerogenes and Enterobacter dissolvens, which did not produce any significant response. The usefulness of the developed immunoassay to detect E. coli in real water samples was also demonstrated.