Characterization of surface properties of glass micropipettes using SEM stereoscopic technique

Abstract

SEM stereoscopic technique is used in this research to determine the three dimensional surface structures of pipettes. Surface properties of a pipette, both at the tip and at the inner wall, play an important role in experiments involving direct contact between a pipette and a biological sample. Tip surface properties of pipettes of different sizes were measured and it was found that the smaller pipette has smaller surface roughness. The inner surface properties of pipette are also measured by cutting half of the pipette using focused ion beam milling. The results show that average surface roughness of pipette tip is 42.2 and 8.3nm for pipettes with tip diameters 8.8 and 1.1μm, respectively. Pipette inner wall has the average surface roughness of 39.0nm and 28.4nm for pipettes with tip diameters 13 and 9μm. The study takes patch clamping as an example. The result of this work can be used to explain one of the major sources of leakage in giga-seal formation in patch clamping. Ions can escape through liquid between the valleys and the cell membrane. The results suggest that as long as the membrane and pipette surface are close enough, the length of the contact is in the second order of importance. This is in good agreement with the practical knowledge in patch clamping that the smaller pipette makes a better seal. The fact that both the tip and the inner wall are rough may promote future FEA analysis and experiments which involve membrane and glass interactions.