Several species of darters of the genus Etheostoma possess a chemical alarm substance, but it is not known how widespread this substance is within the genus. We performed behavioral bioassays on Fantail Darters (E. flabellare), Rainbow Darters (E. caeruleum), and Redfin Darters (E. whipplei). Redfin Darters reacted with a fright response upon exposure to skin extracts from conspecifics and Rainbow Darters. Rainbow Darters, which are known to respond to conspecific extracts, responded to skin extracts from Redfin Darters. Neither Rainbow Darters nor Redfin Darters showed a significant response to Fantail Darter extract, and Fantail Darters did not respond to skin extracts from either congeners or conspecifics. Percid alarm substance is hypothesized to originate from epidermal sacciform cells. Histological examination of epidermal tissue demonstrated that sacciform cells were present in all three species. Our results demonstrate that chemical alarm signals are not ubiquitous for the genus Etheostoma, and that the presence of sacciform cells does not always predict the presence of the alarm response.