I am working on electrochemical biosensor, which measures change of charge transfer resistant for detection of protein. My problem is that the charge transfer resistant keeps changing even if the measurements are repeated on the same electrodes, i. e., after antigen is captured by antibody on working electrodes, multiple EIS measurements did not give consistent results, but kept varying significantly. [My protocol of building the sensor is: attaching antibody to gold working electrode using self-assembled monolayers, using K3Fe(CN)6 and K4Fe(CN)6 as redox probes and doing EIS measurement at formal potenital (about 0.2V) with AC (10 mv from 10kHz to 0.1Hz).]
Does anyone have the same problem before or happen to know the reason for my problem?
Reproducibility of EIS data from SAM monolayer depends on several issues. First, monolayer quality. Second, redox probe concentration. Three, redox probes diffusion through monolayer defects. Collectively, you need to control all these simultaneously.
Finnaly, biomolecular interaction can be active only in certain range of frequency.
Large number literatures dealt with few mwasurements. Ferequent measurement may deteriorate your sensor activity.
Try to check all theses
Dear Dan Wu,
In a study, published in 2002, it was found that light and hexacyanoferrate would attack the gold. The reference is :
M. Dijksma, B. A. Boukamp, B. Kamp, and W. P. van Bennekom, 'Effect of Hexacyanoferrate(II/III) on Self-Assembled Monolayers of Thioctic Acid and 11-Mercaptoundecanoic Acid on Gold', Langmuir 2002, 18, 3105-3112.
The problem is that EIS is very sensitive to changes in the system under study.
I hope thatt his will help.
Best regards,
Bernard A. Boukamp
Dear Dan
I agree with the fact that chloride is not indicated to use with gold electrodes. Also light and hexacyanoferrate are not the best conditions. I found these only after publishing my data. But I also know that a lot of people are publishing with these conditions. You could try to use classical Re or Os based redox probes. If you want to use hexacyanoferrate try to decrease its concentration.
I have also experienced the same problem as you even on bare Au electrodes. The point is that if you could see a trend with the increase of the antigen concentration in your data that is fine enough, because it it really hard to get very good reproducibility.
Good luck.
Is it sensitive enough for your system by measuring EIS but without using redox probe? In principle, it is possible the impedance will increase upon binding of antigens. The ET reaction of K3Fe(CN)6/K4Fe(CN)6 couple sensitively depends on both surface structure and charge state of working electrodes. If both the surface structure and charge state of your systems is in constantly changing, it is hardly expected to get reproducible results.
Still have the problem. 1) I tried to run the measurement in the dark or using chloride-free electrolyte or condition the working electrode in measurement solution for half an hour, still have the problem; 2) I also tried to measure the impedance without redox probe even on the simpliest surface ( thiol peg biotin SAM on working electrode) and I found the impedance kept changing for consecutive measurements, it may become stable after 10s of measurement, but I am afraid that the surface has been change a lot by the applied 200mv dc potential.
Hi Dan,
I am having the same problem with bare electrode. Did you solve the problem?
Hi!
I think many people have the same problem... One nice answer is in this great article; http://pubs.acs.org/doi/abs/10.1021/acs.analchem.5b04814?journalCode=ancham
http://pubs.acs.org/doi/abs/10.1021/acs.analchem.5b04814?journalCode=ancham
I am getting the same problem with carbon based SPE with 5mM K3Fe(CN)6 and K4Fe(CN)6 in DI water as redox probe. If anyone of you have solved the problem, please let me know. Thanks
I am having the same challenge in the laboratory right now. I just saw these various answers tonight. I will try all tomorrow peradventure there will be a change to the inconsistencies I have been experiencing since last week
May I ask how do you prepare your set up in general to have a reproducible result. I also having difficulty getting a consistent result even just using glassy carbon electrode. Thank you
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