This work assesses the feasibility of fluorescence detection of tetracycline in very optically dense mixtures of highly fluorescent media ingredients used in tetracycline production by fermentation. The fluorescence measurements are accomplished with a fiber optic probe. Seven different mixtures were examined in this study. Each one contained a nonfluorescent base of nutrients and salts along with one of the following media ingredients at 5 g/100 mL: cottonseed flour, corn gluten meal, soybean flour, distiller 's grains and solubles, corn steep liquor, brewer's yeast, and molasses. The concentration of tetracycline was varied in each mixture and fluorescence measurements were made at every concentration step. Excitation light of 390 nm was used to probe the samples, and emission spectra were obtained over the wavelength range from 400 to 600 nm. In most of the samples studied, the fluorescence intensity in the wavelength range corresponding to background media fluorescence (420–480 nm) decreased as the tetracycline concentration increased. The decreases in the short wavelength range might be explained by the absorption by tetracycline of 390 nm excitation light (in competition with absorption by the media) and/or by absorption of background media fluorescence by tetracycline. Frequently, the maximum emission of the mixtures shifted to longer wavelengths. The maximum approached that of tetracycline (approximately 520 nm). Plots of integrated fluorescence intensity, in the emission wavelength regions of 420-480 nm and 500-560 nm, versus tetracycline hydrochloride concentration reflect these shifts. We have found that the changes in fluorescence intensity in these two wavelength regions during tetracycline addition depend on the identity of the media component in the mixture. For corn meal, soybean, brewer's, and molasses media, the fluorescence in the short emission wavelength range decreases while that in the long region increases. In the case of distiller's and corn steep media, the fluorescence changes very little during tetracycline addition. Finally, in cottonseed medium, the fluorescence increases in both wavelength ranges. The data show that fluorescence can be used to detect tetracycline, at least qualitatively, in the presence of the highly fluorescent media ingredients.