Two aqueous exposures of fathead minnows (Pimephales promelas) were conducted to determine the bioconcentration factor (BCF) for [14C]octamethylcyclotetrasiloxane (OMCTS). The high volatility, high sorptive capacity, and low water solubility of OMCTS required that the testing be conducted in a novel, totally enclosed, flow‐through exposure system that prevented the volatilization of the test substance. In a preliminary study, fish were continuously exposed to OMCTS at a concentration of approximately 0.5 μg/L for 6 d. In the definitive study, fish were exposed for 28 d at approximately 0.3 μg/L. In each case, OMCTS exposures were followed by a 14‐d depuration period. Water concentrations and fish residues were measured periodically in both studies. Data from the 6‐d study resulted in a maximum measured BCF of 7,000. Model calculations based on data from the 6‐d exposure phase predicted a steady‐state BCF of 10,000 after 19 d of exposure. The definitive study resulted in a steady‐state measured BCF of 12,400 after 28 d of exposure. Model calculations based on the measured data from the 28‐d exposure phase provided an estimated BCF of 13,400. The OMCTS bioconcentration factors could be accurately estimated from the data produced during a 6‐d continuous exposure. The use of innovative test systems was key to the development of useful bioconcentration data.