Hi all, I’m having lately some troubles with my K gluconate internal solution. I made it fresh and freeze it and it works for some days, then after certain time I cannot see any flow (positive pressure) in my patch pipette. I thought at first it was a bad batch of solution, but every time I (or other lab members) make this solution we are having the same issue, working well for some days and then no flow anymore. I checked other possibilities (bad holder, used washer, etc…) but everything works well with ACSF in the pipette or Cs –based solution. I ordered new reagents of every component (except for HEPES) in case something was degraded, but same thing. (I use Kglu, Kcl, Hepes, Sodium Phosphocratine, ATP and GTP, ascorbic acid and NaCl). HELP PLEASE!!!!
K-gluconate is known to precipitate in the solution : you should always sonicate before experiment. You can try to use K-methanesulfonate instead or use larger pipettes.
this will sound really silly, but do you have a filter on your syringe for the internal solution?
Even if the solution is filtered prior to aliquoting, add a filter to the syringe. This always helps when I cannot get positive pressure.
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