The application, control, and removal of cell culture suspensions and experimental compounds are achieved by two open compound addition wells (O1 and O2), a switch valve channel (SW), and a waste well (W). Precise control over the pneumatic pressure applied to O1, SW, and O2 creates a tri-laminar flow with distinct cut-offs between each solution stream in the recording region microchannel.
During the early stages of recording, cells are loaded from well O1 and then control recording buffer perfused to obtain stable baseline recordings (Figure 1A and B). Application of vacuum to SW then disrupts the laminar streams for a time period that can be defined by the operator, and compound is applied from the O2 well (Figure 1C and D). The instrument is able to alter the compound concentration in O2 or exchange to a different compound allowing for multiple recordings from each recording group.
This method of compound application enables precise control over the duration of compound and ligand exposure to cells during recording (Figure 2), with a typical 10–90% solution exchange time of less than 30 ms. This is particularly advantageous when recording from fast ligand-gated ion channels (e.g. nAChR).
More importantly, this also produces tightly regulated reproducible assay conditions to benchmark compound activity throughout medicinal chemistry optimization studies. Table 1 shows the benchmarking of standard compound potencies against literature values, and Table 2 illustrates the reproducibility of assay data obtained using Dynaflow HT over four weeks.
While this first-generation Dynaflow HT instrument is not intended to be used as a full HTS platform, it is particularly well suited to performing smaller, focused screening campaigns against target-directed libraries and will drive the medicinal-chemistry optimization of compound activity/efficacy with greater accuracy by producing high-quality data for each compound evaluated. The initial application data here show the speed and precision necessary as well as the long-term reproducibility required for assaying against all ligand-gated ion channels including fast acting channels.
New technologies have undoubtedly revived pharmaceutical industry interest in ion-channel targets, and Dynaflow HT offers another step forward in the search for an assay format able to really open up the ion-channel field to biologists, medicinal chemists, and the industry as a whole. This renewed opportunity should ensure that the second decade of the 21st century bears the fruit of recent advances and sees an increased volume of new therapeutically relevant ion-channel modulators in development.