I want to do patch-clamping on fish brain slice and I have difficulty to make seals. Osmolarities from both solutions are fine. Dissection is looking really good, my cells seem reasonable healthy. perfusion is just fine. No vibration. Suction is efficient. Kohler illumination verified regularly.
Yet, rarely any seal, no break in. I think my 15 years old infra-red camera doesn't allow a good resolution for proper DIC for electrophysiology. Then, what would be the best camera?
We use the QImaging Rolera XR camera which is a few years old. It works well for IR-DIC and fluorescence. However, if you never get good seals I would imagine your problem lies elsewhere. Even with bad optics you would still be able to get a good seal now and again. The main culprits (as you have identified above) are cell health, internal solution, and electrode size/shape. Smaller electrodes usually make better seals, and internal solutions can be finicky (maybe try a different recipe?). Good luck!
I like cameras from Hamamatsu or Luigs&Neumann, but I agree with Morven, it doesn't look like a camera issue...
The advice of Morven is correct.
In mamalian (rat and mouse) slices, the increase of the glucose extrnal concentration from 10 to 25 mM have been for me helpfull
Try different glass for electrodes change the size of the glass electrode
As recommended by others, I would look at tissue/pipette condition. Would a Live/Dead staining work on fish slice? If the neurons are alive then I would bring some of my pulled patch pipettes to a colleague working on a different tissue (mouse slice?) and ask him to try a few. I hope this helps,
Thomas
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