I have data obtained from patch-clamping on reversal potential of an ion channel. I need to correct my readings for liquid junction potential. I used Axon instruments for path-clamping and I I tried to use the utility in "Clampex" software to calculate the liquid junction potential.
I used 10mM BAPTA ((1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid) in my Pipette solution
However, the database in Clampex do not have BAPTA, therefore I cannot put 10 mM BAPTA in to the liquid junction potential calculation. There is a way to manually enter a new ion/chemical to the Axon data base. However, I need the mobility of BAPTA to do that.
Does anybody know the mobility of BAPTA or a way to calculate the liquid junction potential of solutions contain BAPTA using Clampex software?
Thank You
Ion contribution to the LJP is proportional to ion mobility (along with other factors, of course). Ion mobility is (roughly) inversely proportional to the ion size. According to chemical structure, size of the BAPTA molecule (ion) is substantially larger than that of EGTA, for example. Hence, its mobility is of smaller magnitude than the mobility of EGTA. Most likely, the relative mobility of the BAPTA ion is so small that its contribution to the LJP is negligible, and, for that reason, it is not considered in the Clampex software.
You could measure your LJP directly in current-clamp mode. Initially fill a dish with pipette solution and lower your pipette into it so that the pipette solution is now in both your bath and pipette. Then measure the change in voltage that occurs when you exchange the pipette solution in the dish with normal bathing/extracellular solution.
There is a caveat in the method described by David L Prole. Directly you measure not only an LJP created at the tip of the electrode, but a sum of LJP and the junctional potential (JP) elsewhere including the one at the grounding electrode. JP at the grounding electrode changes with the change of the external solutions. This problem can be obviated by keeping the grounding electrode in an isolated chamber connected to the test chamber with the agar bridge.
Irina is absolutely right - I omitted to mention this as it is often standard procedure to use an agar bridge in ephys recording, as we do (eg. a 3M KCl agar bridge can be used).
Here is a nice protocol that we follow in the lab. It is basically a more elaborate description of the method mentioned by David.
http://junctionpotential.blogspot.de/2010/10/how-do-i-measure-liquid-junction.html
Try to find the mobility of a derivative of Bapta (e.g., Fura-2 for example). I don't know if it is available, but if it is, one can assume that Bapta will have a similar mobility. Having said that, you can experimentally measure it according to the nice protocol given above by Simon. Best...FL-C
Thank you all.
Everyone who are involved in liquid junction potential must refer the page (http://junctionpotential.blogspot.de/2010/10/how-do-i-measure-liquid-junction.html) suggested by Simon. The article and all the comments in that blog are so informative. Specially for the beginners like me.
Again thank you all. This is my first question in Research Gate and it helped me a lot.
Review on liquid junction potentials.
Neher E., Correction for liquid junction potentials in patch clamp experiments, Methods Enzymol. 1992;207:123-31.
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