While a single subject/animal is perfused with multiple concentrations during the no-net-flux method, multiple subjects are perfused with a single concentration during the dynamic no-net-flux (DNNF) method. Data from the different subjects/animals is then combined at each time point for regression analysis allowing determination of the recovery over time. The design of the DNNF calibration method has proven very useful for studies that evaluate the response of endogenous compounds, such as neurotransmitters, to drug challenge.