Pipettes for in vitro whole cell

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vilsy
vilsy's picture
Pipettes for in vitro whole cell

Hi guys,
 
I am currently using 1mm OD and 0.5mm ID borosilicate glass for pulling pipettes for my experiments. I am getting a resistance of about 8-9 MOhms with my current protocol.
I am having trouble getting patches. Can you guys comment if its the resistance of my electrodes that needs to be changed to get better gigaseals.
 
Thanks a million

Fraser Moss
Fraser Moss's picture
8-9 MOhm seems too high.  You

8-9 MOhm seems too high.  You will have difficulty getting a good whole-cell seal because I expect the tip will be too small. If the tip is too small the forces at the tip can sometimes be too great and you get a leaky whole cel seal before you have managed to reach Gigaohm.  Also, even if you mange to go whole cell the access resistance will be high and you will have a large probability of the tip resealing.
4-5MOhm is more desirable.  After you have pulled the pipettes check the inner diameter at the tip of the pipete is between 1-1.5µm on a microscope/microforge.
Also look at the shape of the glass at the very tip.  Does it look smooth and polished or does it come to a sharp point?  You may want to consider pulling wider 2-3 MOhm pipettes and fire polishing them on a microforge back to 4-5 MOhm to improve your seals.