EIS drifts could be related with the breaking away of the protein in the protein-monolayer electrochemical interfaces. What kind of protein do you use? You can check if the protein coverage keep constant by QCM measurements, but firstly i would do the EIS measurements over the monolayers modified electrodes to rule out other posible causes.

Good luck!

If your indication : "No DC voltage", means VDC = 0V, then

you must, always[1], start a (complicated or new) cell with the parametric value of VDC = OCP[2-4].

1. For a minimum of an (additional) external stress/disturbance to your cell.

2. From the OCP (Open Circuit Potential) value(s) you will start running your 1st EIS measurement. You have to run an OCP, to get an idea, near the Open Circuit Potential.

3. OCP_tutorial http://www.ecochemie.nl/download/NovaTutorials/OCP_tutorial.pdf

4. Using “vs. OCP” for Potentials Set in the Sequence Wizard https://www.gamry.com/support/technical-support/advanced-technical-topics/using-vs-ocp-for-potentials-set-in-the-sequence-wizard/

It is importance to take the OCP long time when run EIS as well check the refer. Impedance .

Regards,

Hello everyone,

I observed, from the Nyquist plot, that the impedance for my system became stable after almost 5 years. Throughout this time, the Rct and Cdl kept on increasing till they saturated in last 30 mins or so (plot attached).

To Dr. Santiago :

I am using human thrombin. The MW is ~ 37 kD, at pH 7.4 the charge is ~ + 6 e.

To Prof. Samaras :

Vdc = 0 with respect to Eocp in my case. I am using a Gamry potentiostat. Could you please explain why it would be beneficial to use Vdc = OCP value ? I read the documents you have cited but could not understand the reason behind using Vdc = OCP.

To Prof. Yousif :

I take the OCP measurement for 300 s before doing an EIS. During the first EIS, the OCP is recorded as + 0.097 mV. The variation is around +/-2 per cent during the measurement. In 2 hours, it became + 0.087 mV with variation +/- 1 per cent. At the end, in about 5 hours, the OCP is +/- 0.079 mV with variation +/- 0.01 per cent.

A general question :

Could it be that the system (mercaptohexadecanoic acid SAM activated by EDC , further treated with human thrombin and then with aptamer) took time to equilibrate in the electrochemical cell and as a result of this the double layer was varying in thickness and this is reflected in the Nyquist plot ?

I only used the small AC voltage required for EIS, and, did not use any DC bias voltage thinking that it can disturb the system.

Kindly reply,

Thanks

Dear Agnivo Gosai,

the open circuit potential (OCP[1,2]) is the potential of the WE (working electrode) relative to the RE (reference electrode : Ag/AgCl), when no over-potential or external[3] current is being applied to the cell. When a new potential is applied relative to RE, the system measures the open circuit potential, before turning on the cell, then (leading in ~ms) applies the potential and a current passes between the WE and the CE (Counter electrode : Pt wire). Setting Vdc=OCP (in your EIS), you measure Z (of the WE) left out any external modification. Your EIS[4] might be like an evaluation of a process close to a bio-film[5] coating (of your WE).

So, start an EIS set : Vdc=OCP, and before long ... you might profit from a better process.

1. Agnivo Gosai :"I ... could not understand the reason behind using Vdc = OCP."

2. OCP_tutorial http://www.ecochemie.nl/download/NovaTutorials/OCP_tutorial.pdf

3. or the internal currents are kept in balance (upon any WE modification (your case), corrosion etc.). Using Vdc=OCP, in corrosion means, that we do not force-promote any direction of the i, so the anodic and cathodic reactions are kept in balance. Also, the OCP might result from different processes' contribution(s).

4. EIS, part 1: Fundamentals of Electrochemical Impedance Spectroscopy http://depa.fquim.unam.mx/labcorr/publicaciones/EISfundamentals1.pdf

5. EIS, part 3: evaluation of organic coatings... http://www.paint.org/documents/2017/08/evaluation-organic-coatings-electrochemical-impedance-spectroscopy-part-3-protocols-testing-coatings-eis.pdf

HI,

If you take 300s not enough this is will cause drifting in sample so, at least take one hour before all meaurment .

Regards,

Thanks to everyone and Prof. Samaras

Did you check if you get the same drifts over the monolayer coated electrodes? It could give you a useful information about your system.

Best wishes,

Alain R.

Hi Gosai,

It will better for you to stabilize the aptamers modified gold electrode first (it may take few hours). After that you can add your proteins.

In your case you are using EDC/NHS chemistry to convert COOH groups to NHS and after that you immoblize your aptamers. But it might left some NHS group on the surfacr which can be hydrolysed with time to give you COO- on the surface which repeal your redox probe (I am not sure if you are using Faradaic or non-Faradaic measurement). Even if you are not using a redox couple [Fe(CN)6]3-/4- the interface (double capacitance layer) can be changed during the hydrolysis. Try to block the remaining NHS group with ethanolamine and check your measurment again.

Thanks Prof. Tlili for the insightful suggestion.

Can you calculate the variation of your Rct after two hours of measurement and check the variation of the OCP as well to get more information. After that take new modified electrode and run the impedance measurement at the normal OCP, then you measure your impedance using the OCP (you got it after 2h). This is measurement it will give you a clear idea whether that drift is coming from OCP drift or something else.

Good luck

Hello, I shall update after using ethanolamine. Thanks for the suggestions.

Dear Agnivo Gosai,

if you could implement a Galvanostatic mode for (G)EIS[1,2] during the interval of your specific major[2] electrode's modifications[3], then you might be ready to get (the near real time) hot information, about the processes of these modifications. After the processes of these modifications, you are ready to run the impedance measurement at the final (steady state[4], Vdc=) OCPsteady state. So you are ready, now, to (re)measure, your well known, (P)EIS[5], implementing this final, and safe EIS measurement, upon exercising for long(er) time with this condition : Vdc = OCPsteady state = OCP t ---> oo .

1. Galvanostatic mode (GEIS) for the impedance measurements[6].

2. see, also, my note in a related (corrosion) case, conveniences from GEIS : https://www.researchgate.net/post/How_to_get_good_curves_in_impedance_measurement_technique

3. and the variation of the OCP vs time t, during[6] the initial ~2 to 5 hours, e.g. t ---> oo, in theory.

4. In systems theory, a system or a process is in a steady state if the (state) variables, which define the behavior of the system, and the processes, are unchanging in time.

5. Potentiostatic mode (PEIS) for the impedance measurements : https://www.gamry.com/application-notes/EIS/eis-potentiostatic-galvanostatic-mode/

6. A Galvanostatic mode (GEIS) is, practically, an OCP free, EIS-mode.

Dear Gosai,

The reason for the increase in Rct and Cdl could possibly be the adsorption of protein or complexes on the Au surface and under applied conditions, the current response during potential perturbation is limited because of surface coverage. I can see from the capacitive response in the intermediate frequency region of your impedance spectra which could be related with electrochemical behavior of the bulk electrolyte near the surface ( particularly in the diffuse region of double layer).

The other critical point in your tests is the relatively larger frequency (0.1Hz) which can be selected as 0.01Hz and /or even lower to get the true kinetic information of your system. At low frequency you can truly estimate the interfacial response of the surface i.e. charge transfer, adsorption, etc. The stable OCP and right selection of AC potential (within linear region of potential-current response) perturbation could be the other factors which must be selected with care.