Enhancing throughput with multiple cell lines per well

著者: Hervor L Olsen, Dorthe Nielsen, Morten Sunesen

The QPatch HTX automated patch clamp technology was developed to 1) increase throughput in ion channel drug screening by parallel operation of 48 multi-hole patch clamp sites, each comprising 10 individual patch clamp holes, in a single measurements site on a QPlate X, and 2) diminish problems with low-expressing cell lines. Thus, parallel recording from 10 cells represents a 10-fold signal amplification, and it increases the success rate at each site substantially. To further increase throughput we explored the possibility of simultaneous recording of a number of ion channel currents. Two or three cell lines, each expressing a specific ion channel, were applied at each site simultaneously. The ion channel currents were separated temporally or pharmacologically by proper choices of voltage protocols or ion channel inhibitors. Using this strategy we were successful in recording currents from specific ion channel populations. This strategy, which ensured the exact same conditions for the cell lines with respect to Ringer solutions, temperature, pH and osmolarity, allowed a doubling or even tripling of the throughput. We conducted a series of QPatch HTX experiments with a combination of ion channels involved in cardiac risk assessment: hERG, Kv1.5, KvLQT1/minK and Nav1.5. Specifically, tests were set up for recordings of 1) IV-relationships and concentration-responses for Kv1.5 and Nav1.5 in parallel by using multiple voltage protocols, and 2) pharmacological properties of specific blockers of hERG, KvLQT1/mink and Nav1.5 in parallel. We present biophysical and pharmacological data obtained with QPatch HTX using multiple voltage protocols and cell lines in combination, and compare them to traditional single-hole data obtained with QPatch HT.