A multi-marker approach is increasingly considered in diagnosis of prostate cancer to minimize the number of unnecessary biopsies [1]. Centrifugo-pneumatic flow control is often the method of choice for multiplexing such assays [2]. However, the number of assays that can be parallelized on such a dissolvable-film (DF) based system is limited by the substantial spread of the burst frequencies [2] owing to manufacturing tolerances. In this work we substantially enhance the definition of actuation frequencies by 75% using a novel design to enable the automation of multi-step / multi-reagent Lab-on-a-Disc (LoaD) platform. We demonstrate a multi-marker prostate cancer immunoassay panel.