Correction of liquid junction potential in voltage clamp

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Arirang
Arirang's picture
Correction of liquid junction potential in voltage clamp

 Hi.
I usually use current clamp.
In this state, of course I corrected the LJP.
However, I have no idea about  voltage clamp mode.
Do I have to correct the LJP?
For example, holding potential = -70mV, LPJ=-10mV, therefore real voltage is -80 (=-70+-10)?
Please help me!!!!!

The FFM
The FFM's picture
I refer you to this

I refer you to this conversation that has previously taken place in these message boards regarding Liquid Junction Potential

http://www.scientistsolutions.com/t6028-liquid+junction+potential.html

In particular read the reference

Correction for liquid junction potentials in patch clamp experiments.
Methods Enzymol. 1992;207:123-31.
Pubmed link : http://www.ncbi.nlm.nih.gov/pubmed/1528115?dopt=abstract

Then if you have more questions we can continue here.

Basically, in whole cell voltage clamp patch remember the equation

VM= VP- VLJP

Where VM  = Membrane potantial, VP = pipette voltage and VLJP = the liquid junction potential

So in your example VM = -70mV - (-10mV)  = -60mV 

Arirang
Arirang's picture
Thank you for the quick reply

Thank you for the quick reply.  ^^
I use Axopatch 200B model for patch clamp.
In voltage clamp mode,  display window indicate holding potential (ex -70mV).
If LJP is -10mV, real membrane potential is -60mV (VM= VP- VLJP). I am right?
I can not understand!!!!!
Please let me know. Thank you.

 

lazy
lazy's picture
You can ge a document

You can ge a document entitled "Liquid junction potential corrections" in google. In this document, you can get the information regarding the lilquid junction potential.

For your information.

Arirang
Arirang's picture
:) Thank you so much!!!!!

:) Thank you so much!!!!!

The FFM
The FFM's picture
This is the direct link to

This is the direct link to the Liquid Junction Potential tutorial from Axon Instruments www.moleculardevices.com/pdfs/pCLAMP_AppNote_LJP_Corrections.pdf

The link below is to the software for calculating LJPs as written by PH Barry at the University of New South Wales

 http://web.med.unsw.edu.au/PHBSoft/LJP_Calculator.htm

If you are using Axon Instruments amplifiers, chances are you are also using pClamp and the LJP calculator is integrated in to the pClamp software.

RE: "If LJP is -10mV, real membrane potential is -60mV (VM= VP- VLJP). I am right?"

Yes you are right

Note in the PDF I linked to at the top of this post, they give the example of Vcmd = -50mV and VLJP = +15.6mV.  So VM  = -65.6mV

Arirang
Arirang's picture
Thank you so much!!!!

Thank you so much!!!!
Your suggested link is good for me ^^

zang
zang's picture
I calculated the LJP and I

I calculated the LJP and I got E(b)-E(p)=+10.
After achieving the WC, I beleive this LJP will be gone. So, When I do the offset, I do not want to offset the LJP.

I am using PatchMaster software. Should I enter +10 mV for LJP or -10 mV?
Thank you.

The FFM
The FFM's picture
It is the preferred method to

It is the preferred method to apply LJP correction after an experiment during the analysis.

A different option preferred by many is to do it at the beginning of the experiment.  This is OK provided there are no solution changes during the experiment and the correction is applied correctly.

The LJP is considered to still exist during whole cell patch mode because the contents of the cell is considered to mix efficiently with the intracellular pipette solution.

zang
zang's picture
I just want to make it sure

I just want to make it sure if I understand FFM's comments correctly. To facilitate, I use the following exmaple:

hERG assay
LJP = +10 mV
protocol (Vp): -80 mV _ +20 mV _ -50 mV _ -80 mV

1) If I do not consider about LJP, actual protocol (Vm) wil be -90 mV _ +10 mV _ -60 mV _ -90 mV.
Is this correct?

2) How to apply LJP correction after an experiment during the analysis? Should I mention about actual protocol instead of expected protocol? For example, the holding potential was -90 mV instead of -80mV?

3) The data obtained at different potentials (expected Vm=-50 mV v.s. actual Vm=-60 mV) might be slightly different. How to apply LJP correction after an experiment?

The LJP is considered to still exist during WC patch mode.

3) I have ignored that. Can I? Or, should I do some for this?

Thanks.
 

The FFM
The FFM's picture
 1) If I do not consider

 1) If I do not consider about LJP, actual protocol (Vm) wil be -90 mV _ +10 mV _ -60 mV _ -90 mV.
Is this correct?

YES

2) How to apply LJP correction after an experiment during the analysis? Should I mention about actual protocol instead of expected protocol? For example, the holding potential was -90 mV instead of -80mV?

YES

3) The data obtained at different potentials (expected Vm=-50 mV v.s. actual Vm=-60 mV) might be slightly different. How to apply LJP correction after an experiment?

You measure the peak current amplitude for each epoch in the protocol and when you plot the data the Vm for the peak current during each epoch will be VP-VLJP  where Vp is the potential that was applied to the pipette tip from the amplifier through the control software.

The LJP is considered to still exist during WC patch mode.

3) I have ignored that. Can I? Or, should I do some for this

As long as you know what the LJP was for your previously analyzed data you can go back and adjust the Vm values in the analysis retrospectively

mutant
mutant's picture
 Dear FFM:

 Dear FFM:

I am doing Current Clamp using HEKA-Pulse. I canceled all voltage by clicking "setup" before making seal. After making seal and break seal, I switch it to "current clamp: mode. I read a voltage of -50 MV. Let up say the liquid junction potential is -12 mV. What is the resting potential of that cell, -50 mV or -62 mV? in other words, do I have to correct for that liquid junction potential?

Thank you in advance for your help.  

yao yu

The FFM
The FFM's picture
Yes, in current clamp you

Yes, in current clamp you have to add the recorded current clamp voltage to the measured/calculated value for the liquid junction potential.

In your example -62 mV would be the resting Vm.

You would apply the correction after you have acquired your raw data.

mutant
mutant's picture
 Dear FFM:

 Dear FFM:

You are great. Thank you so much!!

pikappa
pikappa's picture
I usually use voltage clamp.

I usually use voltage clamp. I make voltage clamp experiment in the whole cell configurations and I always correct the LJP (+15mV) in the voltage mode using clampex.
Immediately after the whole cell configuration I switch in the current mode to see the voltage membrane of the cell.
Do I read the correct Vm or I had to correct it also in current mode?
If so, can I see the true value for Vm if I enter +15mV  in the lab bench dialog box?