The effect of aperture size on gigaseal formation

Malboubi, Majid; Behroozi, Mohammad; Bowen, James; Chirazi, Mahmoud; Charras, Guillaume and Jiang, Kyle (2013). The effect of aperture size on gigaseal formation. In: 57th Annual Meeting of the Biophysical Society, 2-6 Feb 2013, Philadelphia, USA.



Patch clamping, the gold standard for ion channel studies, is entirely dependent on formation of a high resistance seal between cell membrane and patching site,known as gigaseal. As this process is laborious and time consuming, there have been many attempts to develop automated high throughput chip-based patch clamping devices. In spite of recent advances, these devices still cannot form gigaseals relying instead on less tight seals that impede their ability to measure the pA ionic currents passing through single ion channels. Progress is presently limited due to a lack of understanding of the physical and chemical mechanisms underlying gigaseal formation. In all forms of patch clamping access to the cell is achieved via a small aperture. Here, we systematically examine the influence of aperture size, micropipette rim morphology, and surface roughness on gigaseal formation in conventional patch clamping using micro/nanofabrication and modelling techniques. Our results show that smaller aperture sizes lead to improved seal formation within a range of x-y. For aperture sizes out of this range, either bigger or smaller, gigaseal formation is very difficult if not impossible. While in the literature the surface quality of patching sites is only described by average surface roughness, this research reveals that parameters such as: developed interfacial area ratio, valley void volume of the surface, ratio of core void volume to core material volume, and maximum peak to valley distance play more important roles in seal formation. Furthermore,these parameters are size dependent; as a result glass micropipettes with smaller aperture sizes are flatter and have lower water retentionability resulting in better seals. Results of this work support the practical knowledge that pipettes having smaller apertures form better seals.

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