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Electrochemical Biosensors - Science topic

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Hi. I am currenty working on electrochemical biosensors, using 2mM ferro/ferricyanide solution in Pbs and 100mM kcl. I made it carefully and done deaeration with nitrogen gas then Stored in a dark closet in a glass container. But it keeps getting decolored within weeks. I don't know if it is my mistake or it is naturall. My prevoius samples without support electrolyte (kcl) from monthes ago is still strongly colored. Thank you
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First, I want to thank you for your time.
If I understood correctly, your idea about the color change is that a redox reaction occurred in my solutions oxidizing chloride ions to make Cl2 and reducing Fe(CN)6 complex. but how is it possible to happen spontaneously when ferri/ferro cyanide cell (2mM) has a positive Ecell(0.37) and there is no chlorine gas to reduce?
furthermore, both ferri/ ferro cyanide are colored red/yellow. if there is a redox equilibrium the color shouldn't be removed or reduced.
Some of the decolored electrolytes were not even used a single time.
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I am looking for published data on the variation / strength / reach of the electric field around the working electrode for a typical 2/3 electrode electrochemical biosensor. Specifically , I am interested to model the electric field to understand the distance from the electrode where its effect / strength becomes negligible.
I want to understand how this limits the performance of biosensors, specifically those which measure impedance , capacitance etc.
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Anne Sawhney's answer is dead on. Unless you're doing something very unusual the biosensor is likely to be in a fairly high ionic strength environment so most of the field will dissipate by the outer Helmholtz layer (< 1 nm). There will be a diffuse double layer but that is more related to concentration gradients, not field strength.
That said, if the distance between the current carrying electrodes is great enough, the current itself high enough, and the impedance across the gap (tortuous paths) high enough then there can be a field effect away from the electrodes themselves. This is sometimes made use of to induce migration effects (e.g. capillaries). A simple check to figure out if that is the case: if the total voltage dropped [between current carrying electrodes] is about the same as the electrochemical potential then field effects away from the electrodes are minimal. Impedance testing can complicate things but not overly so as it's a small signal. Still the high frequency components do bypass the electrode double layer but they aren't doing anything besides shifting more mobile ions back and forth a little bit out in solution.
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Hi all,
I am really new to electrochemistry and I am recently trying to reproduce the cyclic voltammetry of Pt electrode that I made using sputtering. I used Ag/AgCl as reference, coil Pt as counter and my Pt electrode roughly 3x3 mm2 as the working electrode. The CV graph that I got is quit different from what we usually see on lots of literatures. see the attachment
Could someone please help with this.
Many thanks
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The higher current density means that the surface area of your "Pt" is much higher than the geometric surface area of your electrode. The cleaning and preconditioning of Pt-electrode is tricky. As mentioned by Boris kasrielovich Filanovsky you can use piranha (not pirana) solution.
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i have uploaded image about question.i want to design some biosensor in electrochemistry. i need two chemical molecules (A & B). A is conjugated on a surface by some linker and B is connected to a electrode surface. when the linker breaks, A and B should joint each other especially with targeted linkage. A should have a property to induce electric current in B.
please guide me which molecules can be A and B?
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Good afternoon, you can decorated many electroactive lable by streptavidin, e.g. nanoparticles, HRP enzymes or..., there are also many commercial electrochemical lables based on Streptavidin.
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During the electrochemical analysis of the electrodes, I want to know the parameters of CV , DPV , EIS and how to add the concentration of analyte in the electrolyte. I am using PBS as an electrolyte and GCE as the working electrode.
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As you commented, there are a wide variety of electrochemical techniques. Based on the type of biosensor (affinity of biocatalytic). As affinity biosensors operate via selective binding between the analyte and the biological component, voltammetric and EIS are common techniques used. Biocatalytic biosensors incorporate biomolecules such as enzymes that recognize the target analyte, and subsequently produce an electroactive species, and thus amperometric and voltammetric techniques are generally used.
The electrochemical parameters are based on your detection system (e.g., potential of redox probe, potential of catalytic process, etc.). In terms of how to add analyte in the electrolyte, it will depend on what approach you use for the development of your biosensor. If you develop an enzymatic biosensor, amperometric i-t curve is very useful.
My suggestions would be to identify a few similar related papers to your project and to determine the ideal conditions.
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We have designed a dual enzyme electrochemical biosensor that generates glucose at the end and ultimately to H2O2 to generate the signal with help of glucose oxidase. We would like to challenge the proposed biosensor in serum samples however, glucose that is already presented in those samples in high concentrations I suppose can interfere with the signal. Can someone suggest how to avoid the interference?
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Hi, maybe you should check the amount of glucose in serum by a YSI analyzer first.
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I am trying to run cyclic voltammetry experiments on a modified electrode. The literature shows distinct peaks for the analytes (pesticides, phenols) using glassy carbon electrode(GCE) as working electrode. I am able to get the peak signal on GCE but not on the modified electrode. What could be the possible reasons?
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with great pleasure my dear
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I'm working on electrochemical sensing drug molecule. My workstation is Origalys (origaflex 500). In papersa particular concentration is reported for every sensing studies like 200micromolar etc. How to do chronoamperometry for the sensing application and what is the relevance of this concentration. How to get this value of concentration
Please share your ideas
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Interesting query
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I am currently determining EIS spectra on different carbon-based electrodes. For this, I use a PalmsensPS4 at pretty standard conditions in a three electrode setup (0.1 Hz to 100 kHz) and measure against OCP in a 5mM [Fe(CN6)]3- / [Fe(CN6)]4- (1:1) in phosphate buffered saline (PBS) at pH 7. I then use EIS Spectrum analyser to fit the curves. One type of my electrodes results in data that can be easily fitted with a standard electrochemical Randles cell (yellow in figure). The other type, however, seems to have lower impedance overall, and very low charge-transfer resistance in particular, and I cannot seem to find a model that fits it (blue in figure). When I modify the electrodes with PEDOT, I get the same picture.
I have some questions relating to this:
(1) How can I explain, in EIS terms, what happened to these electrodes?
(2) Which model can I use to generate useful fits?
(3) Are there any papers in which these strange EIS curves have been obtained and are explained?
(4) Is it worth to change experimental conditions and what would you change to obtain meaningful data? I was planning to change the electrolyte concentration and / or the potential (though I am not sure if that would give any meaningful results).
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Don't mention it. We're happy to help. Like the medievals said, "respondeo dicendum":
(1) You can see your Bode plot (the -Phase one) with some peaks in this plot. I've attached here your plot with the max points I've told you earlier. So, in both experiments you have three maximum points in phase angle: in high frequencies (circa 105 Hz, which is the double layer phenomena in the electrode/surface region), another one in middle frequencies (circa 103 Hz, which is the charge transfer reaction), and another one in lower frequencies (between 100 Hz and 10-1 Hz, which is the electrochemical diffusional region). Since then, you have some interesting things here: now you can undersand better which goes on your electrode's surface, since there you have two behaviors: at high frequency the blue plot shows a very tiny semicapacitive circle (as seen in Nyquist's plot) but vanishes in the yellowed plot; the middle frequencies increased the phase angle (which means a decrease of resistive behavior of your electrode's surface), and finally in lower frequencies the diffusion changes between blue and yellow. So, maybe your modification improved the performance of ferri/ferro electrochemical reaction. Being a SPE electrode, the mass transfers changes a lot. For instance, the diffusion changes for a "semi-infinite" to a finite diffusion due to the diminshing surface area of your electrode. So, the classical Randles diffusional electrochemical circuit won't apply in your case and everything in your system looks fine.
(2) Yes, that's OK. The KK test isn't so easy to understand but their results are. See, your test showed a residual error less than 3% in all frequency range, and your KK data shows which I assumed in my answer: your system looks good.
(3) Yeah, impedance is beautiful but the current literature lacks on those experimental things. But I really like to discuss those electrochemical things in my research papers, and maybe sometime I'll publish some literature regarding this.
Hope it helps, and fell free to discuss anytime. 🤓
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Hello, I am doing a carbon surface modification with Gold nanoparticles.
from the FTIR below, I know the formation bond of NH3 and for COO , but I would like to know what is the range of thiol bond formation, is this FTIR good or not?
note that I know that the difference between the bare SPE and modified SPE is the increase of current which is measured by Potentiostat.
thanks in advance.
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Dear Aseel Alnaimi, the above FTIR spectrum is not appropriate given the fact that the corresponding transmission percentages of all peaks are higher than 90% signifying that the sample content in your KBr pellete has been rather too low. In such a condition, on account of low signal to noise ratio, some characteristic peaks are embedded in the noise level and cannot be detected at all. So, I suggest you either repeate the FTIR analysis once more with a well-prepared KBr pellete or use ATR-FTIR technique which can provide you with a higher S/N ratio for the surface functional groups. However, FTIR spectrum might not solely justify what you are dealing with but if it is accompanied with other surface characterizations especially XPS, it will most likely lead to a more rational conclusion. Please note, the characteristic peaks of the carbon materials are usually of low intensity, that is why it is mostly preferred to use XPS as a more reliable technique for inspecting their chemical structure.
Best,
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I am working with screen printed carbon electrode and FTO coated glass electrodes for sensor studies. My electrolyte is PBS solution (pH=7). Please discuss what are the initial and final frequency that are commonly used for recording EIS spectra of these electrodes (bare electrodes).
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There is a very simple way to determine the frequency range in which to perform research - this is the frequency range of the measuring setup.
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Recently, I am working on a new project for developing the glucose biosensor based on electrochemical measurement. I don't have too much experience in electrochemical analysis and electrochemical biosensors, so I would like to ask some questions and I hope some experienced researcher can help me out.
1. I read several papers related to the glucose biosensor and in the papers, most of them were using the Prussian Blue as the mediator for transferring the electron from the enzyme (glucose oxidase) to the electrode. However, the synthesis of the Prussian Blue required dissolving the K3[Fe(CN)6] in the hydrogen chloride solution ( which is a strongly acidic solution). And I am worried about is there any risk of mixing the K3[Fe(CN)6] with HCl? For example, is there any possibility that there will be the release of HCN?
2. I want to use chronoamperometry for the measurement but I don't know how to select the appropriate potential for the measurement? I think I will do the measurement with the selected potential and based on the current change, I can determine the glucose concentration. So should I do the CV firstly and then which potential should I choose? Potential of peak current in the CV or others?
Thank you so much for the reading. And please give me some ideas. Thank you in advance.
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1 Don't worry about the addition
2 You can use the CVs to determine the oxidation potentians for you substance first. Then use the DPV or SWV to get the oxidation peak current values. CVs is the most reliable techniqe for potentials analysis and the SWV or DPV has lower background currents, which are more accurate in peak currents than CVs. You can read the book below. Also a latest work finished by our team took the strategy above, just for reference.
ELECTROCHEMICAL METHODS
Fundamentals and Applications. Allen J. Bard
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I am using screen printed carbon electrode (scpe) for cyclic voltammetry in my sensing studies. The electrolyte is PBS solution (pH=7). I got the CV for bare SCPE without much noise but after coating of the nanomaterial I couldnt get any proper response instead very high noise in the CV. I tried with two different nanomaterials and tried two different coating methods (without and with binder), but the observation is same.
I am attaching the picture of noise I got.
Please discuss your valuable suggestions for solving this issue. I haven't done any pretreatment for the scpe before coating the sample
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In electrohemistry, noise ideally needs to be removed t source since electrochemical devices are typically non-linear for most of the i-V characteristics and noise will be rectified leading to current offsets. It is possible that it is simply a coincidence that the nanoparticle modification happened before the noise increased. Do a quick Fourier transform and see what the dominant frequencies are- this could let you know if it is pick-up from the power supply- as this will be 50 Hz (UK and Europe, or 60 Hz US) and probably the odd harmonics, but make sure that the sampling frequency is as high as possible, but at least >100 Hz. Did you change the sampling frequency after the treatment? Your noise might previously have been aliased to very low frequencies and not seen. Also, what reference electrode are you using? If this is the one on the screen printed electrode, these are pretty terrible at the best of times, but some alteration in the surface chemistry could lead to increase Ref-solution impedance. Changes in contact resistance after the treatment is another possibility.There are some useful tips on noise in our paper "Lifting the lid on the potentiostat" and the electronic supplementary information shows real world examples: Physical Chemistry Chemical Physics (2021) 23 8300.
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I design an electrochemical biosensor for the early diagnosis of breast cancer. I must determine the -NH2-ended aptamer to catch HER-2. I haven't decided yet, I would like to ask you. I will analyze using electrochemical methods such as CV, EIS or DPV whether the aptamer is bound to the target HER-2 or not.
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Dear Zeynep Turk , That depends upon pH of system to observe the -NH2 in electrochemical process.
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When I researched articles on the electrochemical applications of nano-Au.
I found this figure and noticed an extra single cycle. And when I did my Nano Au CV analysis, I didn't get that extra single cycle. So, what is that cycle? Why does it appear on the cyclic voltammetry?
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Yeah, I just gone the the figure in that article. What I understand is that in Figure A they are doing electrochemical reduction of graphene oxide (GO) to reduce graphene oxide (rGO) using cyclic voltammetry. at first cycle reduction current is very high as more GO get reduce in1st cycle, then reduction current decreases with more number of cycling as there are less GO left in each cycle.
In the Figure B similar reduction process happened . Here "HAuCl4" salt(Au3+) get reduce to Au on rGO/GCE surface. And reduction current is very high in 1st cycle, then decrease with cycling.
Hope you got the answer!
Best regards,
Bapi
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I want to keep my pencil graphite electrodes for a while in the aqueous graphene oxide solution that I have prepared at different concentrations such as 5 mg/mL, 1 mg/mL, 0.5 mg/mL, and then i would like to make graphene oxide adsorbed on the surface of PGEs and then i will reduce the graphene oxide electrochemically. After a while, the solution becomes heterogeneous as it begins to precipitate. Have you ever encountered such a situation before? It sounds good to dissolve it in a phosphate or acetate buffer solutions, in case this may related with pH, has anyone experienced it?
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  • Add filesZeynep Turk , if your GO dispersion is per se stable and does not precipitate spontaneously, it most likely originates from the hydroxide ions produced on the cathode in electroreduction process. GO nanosheets tend to precipitate upon introduction of hydroxide ions as result of removal of the oxidation debris attached on their basal plane. In fact, during the synthesis procedure some highly oxidized polycyclic aromatic hydrocarbons are formed which attach to the GO and make it highly hydrophilic. However, these species are detached at high pH values and GO nanosheets precipitate immediately. So, if you are confident that your GO is highly hydrophilic and the prepaired dispersion is stable over time, I suggest you use a proper buffer solution with low ionic strength. Note, high ionic strength can precipitate GO as well, so try to use a dilute buffer with nearly acidic pH. Not to mention, if I were you, I would drop cast and dry GO disperion on the surface of the pencil graphite and then heat it for a while in an oven at 80 degrees. Afterward. I would sufficiently reduce the bonded GO using several successive cyclic voltommetry cycles. By the way, if you merely want to deposit rGO on the pencil electrode and the electroreduction does not matter to you, it is more convenient to drop cast GO and then reduce it at high tempratures. Anyway, as I stated above, it is probable the problem arises from your GO itself. So if your dispersion does not look like the attached photo, I suggest you synthesize GO again. I have optimized and reported the GO synthesis conditions by improved hummers method in the following paper. It may help you to get a high quality GO. Please note, the drying conditions reported in the article is a determinant step and should not be ignored.
Best,
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m looking for a definition of a term or phrase
I have a different research-related question
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I don't know how to consider the design of the electrochemical biosensor based on chitosan and poly aniline. If you mean laboratory design. The first search to do is look for has any one ever done such research if done you can read a few articles about this topic and get information . If no one has done it yet and you want to do it First. In the laboratory method; biosensor design in article can be based on any material and you can use the method of that article to design the bio sensor that you want but at this time you have to change the material on which the biosensor is designed and insert chitosan and poly aniline and if you want to do design using simulation soft ware such as com sole and hysys you must find the same article that designed the biosensor with laboratory method and use the laboratory data of the article and simulate the biosensor according to the article and compare the results with laboratory results and get the error percentage and then change the material which the sensor is designed based on and put chitosan and poly aniline then get the result . These definition that explained was about the bio sensor designing. Now if you're question is about biosensor and poly aniline and chitosan should be discussed separately
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Hi All,
Did anybody encounter a situation when using a planar electrochemical biosensor to detect O2 results in a significantly weaker current (~20% lower) for whole blood compared to that of H2O? Both whole blood and water are equilibrated to the ambient, so, theoretically should provide the same I.
Thank you,
Alex.
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Could you give more details on your O2 biosensor? I might simply come from the difference of media. Whole blood is generally more viscous than water and contains red and white cells, as well as platelets. Therefore, the diffusion of oxygen to the electrode surface could be delayed comparing to H2O.
Best regards,
Marie B.
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I am trying to measure resistance from I-V curve from a chemiresistor/ FET sensor with a keithly sourcemeter. The substrate is soaked/ wet with protein and phosphate buffer only. But the I-V line is not passing through the origin (which probably means it's not showing ohmic characteristics), rather its going through the second quadrant. What could be the possible reason. Picture is attached for reference. Thanks in advance.
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maybe its an offset, check your device on a dummy cell
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my working electrode is graphite sheet and I'm using Ag/AgCl and pt electrode as reference and counter. I have problem with my solution. in some articles, they have used PBS + KCl solution and they observe the redox peaks. in some other works, they use PBS + KCl + K3[Fe(CN)]6 and they see peaks as well.
I already use PBS + KCl solution but I didn't see any peaks for redox reactions and I have a lot of noise in my CV curve.
I don't know what solution is gonna work for me.
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In the absence of the redox active ferro/ferricyanide, you would not expect any peaks in the CV unless (i) dissolved O2 has not been removed and the potential range includes negative excursions or (ii) the graphite had been oxidised and had quinone groups. The potential limits, the range in which no voltammetric features are observed (because nothing is being oxidised or reduced) depends on the pH, but potentials more positive than ca. +0.8 V to Ag/AgCl/ 3M KCl could oxidise the graphite and >1.0 V or so could electrolyse the water to O2 and protons.
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I think the methods that have been used in
Electrochemical Biosensors like ( antibodies- aptamers-MIPs) are usually used for the short-term as we need to wash the electrode to remove the targets from it. I think it is a drawback if we want to use them in daily life wearables such as ( smart wrist band) as the user needs to change the electrode or wash it after a while of usage.
Any suggestion?
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Hi,
I'm not currently Involved in this myself, but the following two very recent review articles (definitely the first one) pay attention to this topic. The first one is focused on implantable sensors, the second one for various monitoring applications.
Kind regards,
Mats
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For a range of concentrations of protein, I am getting the I-V (Current-voltage) data from a Keithley sourcemeter. Accordingly, I am getting the resistance as the response of protein-protein binding on carbon nanotube on my biosensor. What formula should I use to get normalized response versus concentration gradient? Is there any good textbook on this to show the calculations step by step from scratch? Thanks in advance.
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Dear Touhid,
Prof. Reg Penner's work in Nano Lett. with virus bioresistors for HSA protein detection may be of interest to you. Figure 5 shows the change in resistance as a function of protein concentration (calibration curve). The mathematical fits to the Hill equation are outlined in detail, with more information provided in the Supporting Information. EIS data is also provided to establish an equivalent circuit. This paper may represent a good model for comparison as you put your work together!
Kind Regards,
Matt Glasscott
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During doing cyclic voltammetry in three electrodes system (reference electrode, contour or auxiliary electrode, and the Au working electrode), tiny bubbles form at the surface of the Au working electrode. Does anyone have any suggestions about how to get rid of bubbles without changing the potential window?
Thank you.
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Is your supporting electrolyte oxygenated or de-oxygenated? In both cases those bubbles could produce a noisy voltammogram. You can get rid of them by purging a bit of the gas that you use for your experiments.
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Hello all,
We are using screen printed gold electrode(220BT) from Dropsens to build an immunosensor.
To attach the antibody onto the working electrode, we use EDC/NHS with MPA.
From what I understand, we need to restrict the linker and antibody adsorption on working electrode alone. However, many of the paper I read that uses the same electrode only mention they "drop 50 ul of liquid" onto the electrode which got me confused.
From what I can see, any liquid over 10ul will spread from the working electrode and cover the reference and counter electrode but I have not seen much mention of flow chamber or whatever else that can be used to contain the liquid to the working electrode.
Is this something that was just omitted because it is too obvious or is it just common to drop the liquid on to the entire electrode?
If there is an obvious way to isolate the working electrode, could you let me know what it is?
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I want to measure the resistance of medium at a fixed frequency by inducing constant ac current and then measuring the voltage drop across the medium. Ag/AgCl pallets are cheaper than Ag/AgCl reference electrodes. What is the actual difference between them and how are they different and what types of experiments are they most suitable for?
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Dear Dr. Bibek Raut,
the use of Ag/AgCl quasi-reference electrodes, can interfere with the electrochemical measurements and cause non-negligible artifacts. The artifacts are attributed to the redox reactions and adsorption/desorption processes of Ag/AgCl residues of quasi-reference electrodes that dissolve in solution on top of a microelectrode array chip. Having performed a range of investigative experiments, we come to a conclusion that these signals come from direct oxidation/reduction of Ag/Ag+ nanoparticles at the microelectrodes.
For more details, please see the source:
-Electrochemical artifacts originating from nanoparticle contamination by Ag/AgCl quasi-reference electrodes
Alexey Yakushenko, Dirk Mayer, Johan Buitenhuis, Andreas Offenhäusser and Bernhard Wolfrum
Lab Chip, 14, 602-607 (2014)
Best regards, Pierluigi Traverso.
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I wnat to develop electrochemical biosensor to detect miRNA.
I have probe miRNA
Target miRNA
single mimatch mirna
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Biomarkers are the target antigen/analytes. If you want to develop electrochemical sensors on AuNPs coated substrate, you have to treat/functionalize your substrate with some thiol groups/-COOH/-CHO/-NH2 groups. These functional groups can covalently immobilize antibodies/oxidase.
For further information, you can look over my articles ....
Thank you.
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Some label free electrochemical biosensors based on affinity events between immobilized antibodies and proteins use the ferricyanide/ferrocyanide redox couple as a redox active probe. These molecules have a negative charge of -3 and -4.
Are there any other redox molecules that perhaps have a positive charge (cation) that can also be used as a probe?
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Hi Riaan
A popular choice in standard electrochemistry is Ruthenium Hexammine. This behaves more ideally than the ferri-/ferrocyanide couple, and you typically see it fouling your electrodes less. It is however noticeably more expensive.
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screen printed carbon electrode was characterized using 5 mM Ferricyanide/ 5mM Ferrocyanide with 0.1 M KCL , with scan rate from 10 to 100 mV/s vs Ag/AgCl , why the shift go to the right then to the left?
the relation between the square root of the scan rate and the delta potential( shift) does not linear?
please find the attached attached figures which are :
1-cyclic voltagram,
2- delta vs square root of the scan rate.
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It's indeed important to consider only steady state voltammograms but from the figure I assume this is the case. The negative shift is not to dramatic, but one explanation could involve the hydration ferrocyanide which may be a limiting factor at the screen printed electrode. Ferro(hex)cyanide strongly coordinates water (about 6 molecules). Consequently you can think of ferrocyanide as complex with water, when there is less water ferrocyanide becomes more "free" and the reduction potential shifts to more negative potentials, as you can verify by including water in the Nernst equation. This can be further demonstrated by using mixed solvents of varying composition (e.g. water/acetonitrile), the redox potential will then shift to more negative values as a function of increasing acetonitrile concentration. So some solvent residues (from the printing), or the hydrophobicity of the electrode can explain this slight negative shift.
Best regards, Mike
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First, i coated Hexanedithiol on to the gold surface and then blocking remain gold surface used Mercaptohexanol. After that Benzoquinone was coated on gold surface. We reduced the benzoquinone using electricity. Finaly, thiolated peptide was coated on to the gold. And measured using SWV.
(Buffer-4mM Fe(CN)3-/4- )
Question 1. We can't observed clean peak in SWV. How can i get a clean peak?
Question 2. Some research papers suggest that SWV should measured many times to get stable and clean peaks. Is this suggest
correct?
Thank you for interest to my questions.
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Chaehwan Cho check your frequency. You need to run your experiment on different frequency e.g. 5, 10 or 15 and choose the one that is very clear. I am very much sure you will resolved your issue and will get very clear results.
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I have seen many researchers including Prof Alan Bond measure the effective areas by measurement of the peak current obtained as a function of scan rate under linear sweep voltammetric conditions for the one-electron oxidation of Fc [1.0 mM in CH3CN (0.1 M Bu4NPF6)] or reduction of [Fe(CN)6] 3- [1.0 mM in water (0.5 M KCl)] and use of the Randles-Sevcik equation.
The classic IUPAC paper (Trasatti and Petrii 1991, https://www.sciencedirect.com/science/article/pii/002207289280162W) provided us with some other important recommendations in different conditions (one example is via reduction of gold oxide)!
I would like to know whether there are any disadvantages of using the Randles-Sevcik equation for measuring the area? Do you prefer any other methods over this one while working with a reversible reaction on gold disk electrode?
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Firstly, there is an important distinction to be made between the “surface area” and “geometric area” of an electrode. Surface specific methods, such as gold oxide stripping, will tell you about the former, as pointed out by Danny O'Hare , in his response. Redox mediator methods, such as the oxidation of Fc or reduction of [Fe(CN)6]3- will tell you about the latter. This is because even an atomically rough surface will look “flat” on the scale of the diffusion layer, hence the diffusion limited current is sensitive to the geometric area, rather than the true surface area (note that the ratio between the two, surface area/geometric area, is the “roughness factor” of your electrode).
As for your original question, while it is true that Prof Bond often uses the Randles-Sevcik Method to calibrate the geometric area of electrodes used in electroanalysis, two important assumptions need to be made in this case:
1) The reaction is electrochemically reversible (Nernstian) on the timescale of the experiment –this is always true for Fc on metal electrodes at scan rates less than 1 V/s, but may not be true for the case of [Fe(CN)6]3-, which can exhibit some deviation from reversibility on some electrodes.
2) The contribution from uncompensated resistance, that is iRu, is negligible.
The voltammetric peak current, ip, is influence by both the electron-transfer kinetics (k0) and iRu, making it a less than ideal form of analysis.
Another method that you could try is semiintegration, which converts your peak-shaped transient voltammogram into a sigmoidal-shaped semiintegral voltammogram. Unlike the peak current, the diffusion-limited plateau of a semiintegral voltammogram is insensitive to both k0 and iRu.
Details can be found in: Bentley, C. L.; Bond, A. M.; Hollenkamp, A. F.; Mahon, P. J.; Zhang, J., Advantages available in the application of the semi-integral electroanalysis technique for the determination of diffusion coefficients in the highly viscous ionic liquid 1-methyl-3-octylimidazolium hexafluorophosphate. Anal Chem 2013, 85 (4), 2239-45.
Hope this helps,
Cameron Bentley
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I have gold nanoparticles suspension in citrate buffer and i want to modify the WE of screen printed electrode with is particles using thiosalicylic acid , if any one faced similar work, please give me guidelines or the protocol was used.
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Hi all,
Am working on a DNA based biosensor, DNA are thiolated. i reduce thiolated ssDNA by TCEP and dip cast it on a gold nanoparticle coated electrode.
I have tried with 500nM , 1uM, 2uM, 3uM, 4uM and 5uM concentrations, but am not getting a constant value by DPV.
i use methylene blue as indicator, should i have to refrigerate the methylene blue after use.
i need to fix the probe concentration.
Can some one share their insights on this, that would be great help.
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Dear Hammond,
The oligo am using is Thiol C6-Modified, The electrodes are so small that, it can be dipped in 96 Well plates. Am adding 150ul of Oligo's of different concentrations to the well (the volume is made such that the working electrode has to be completely submerged) and dipped the electrode for 180 mins at 30o C. Washed with buffer to remove any unbound oligo's and treated with 6-Mercapto-1-hexanol (MCH) for an hour at 30o C, further washed with buffer to remove any unbound MCH and electrochemical studies are carried out.
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i have carbon screen printed electrode and gold nano particles suspension in citrate buffer, how to deposit the particles on the electrode in a way to get uniform distribution?
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Masoomeh Ashoorirad any recommendation?
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I am looking into electrochemical biosensors based on square wave voltammetry and cyclic voltammetry. These sensors usually use potassium ferricyanide/ferrocyanide as the electroactive species in an electrolytic solution such as phosphate buffer saline (PBS). Most articles site that a concentration of 5 mM potassium ferricyanide in PBS is used for conducting voltammetry experiments, but I would like to increase this to about 10 mM in PBS to improve my signal to noise ratio.
I was wondering what effect this would have on the proteins, such as antibodies, on the surface of the sensor? Could denaturation of antibodies and target proteins occur when the concentration is too high?
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Thank you Falk Fish and Azam Farshadinia for your reply.
In response to your question, I personally do not think that 10 mM would be too high considering that you are using a transduction method like CV. Concerning the effect on biomolecules (antigen, antibodies, proteins or else...), it should not be a problem if you are working in mM. Higher concentration like 0,1M or higher could potentially be an issue and like Falk Fish stated earlier you could probably just make a few tests to see that. Although if you are working with very expensive biomolecules this could be an issue as well...
However, I have to say that the use of a buffer is very important when conducting electrochemical experiments on biosensors since the denaturation of biomolecules could occur if the buffer that you are using do not provide the necessary conditions (pH, concentration, temperature, nature of the electrolyte ...) for their optimal activity.
Finally, if you would want to use impedance for example, increasing the redox probe concentration would drastically change the shape of the spectra, but this should not affect the biomolecules.
Best regards,
Marie B.
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How to choose the target DNA samples for electrochemical biosensors?
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Hello Vinoth Raja ,
Do you have a specific application in mind or are you simply trying to perform a rapid test to confirm behaviour?
There are many suppliers of oligonucleotides and you can often design your own sequences.
Of course, for the probe you often want an anchoring group, such as a thiol (-SH) followed by a spacer sequence (such as PEG) to distance the label sequence from the anchor and sometimes to add flexibility. As for the label of the target sequence, it is often wise to purchase a series of mismatches (mismatch in the centre, and at the 5' and 3' positions) as well as the complete mismatch sequence.
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hello all ,
in my lab i have PGZ100( EIS Maximum frequency 100 kHz , Best current resolution 30 pA, Scan rate 0.5 V/s , Lowest current range 1 µA , Frequencies/decade 5,10,15)
does this potentiostat suitable for biosensor characterization? and how i can judge if it good for my work or i have to use another potenntiostat?
thank you
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I agree with Marie Berthuel, but do want to emphasize that while the low current range is the most likely specifcation that is going to limit you, you should be very careful with EIS measurements if you are going to be anywhere close to the system limits.
I would strongly recommend running some EIS tests on dummy cells to establish how well the system performs. In your case probably a 10 kOhm resistor--maybe larger if your electrodes are small--and an open lead [EIS] test. It would probably also be useful to run a (10?) nF capacitor.
If you get good data from the resistor and capacitor you can have better confidence in the data you are getting from your sample.
The open lead test is to establish a hardware limit. It won't be great but based on the specs it should be better than 1 MOhm at low frequencies.
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Currently Iam working on amperometric biosensor using SPCE (screen printed carbon electrode) but I still confuse to figure out what is the different between this electrode with Interdigitated array electrode (IDA) since I want to know whether I can work at the same condition between these two. 
Thank you.
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Hi Tasneem,
A screen-printed electrode (SPE) refers to the way of fabrication, and an IDA electrode refers to the geometry (two interdigitated series). Usually IDAs are fabricated following a thin-film technology (nm-scale layers) and SPEs form thick films (micrometer-scale layers). The main advantage of IDAs is redox cycling, then, the choice will depend on the system.
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hello!
iam working on a biosensor design, to detect her2 protein using single strand DNA BIORECEPTOR, i need buffer to unbind thiol-modified oligo from AuNP, also a buffer to unbind PROTEIN-DNA ?
please if any one have an idea let us know!
thanks
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Removing Thiol ligands from Au surfaces is quite difficult without using another thiol based agent -> which would render the AuNP useless for further binding studies.
So my elaborate guess would be: use something like a diamine in high concentration and see if it works (ethylendiamine is the weapon of choice id say in this case).
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Dear Researchers/Scientists,
I have recorded the CVs at different scan rate (100-1000 mV) and plotted the graphs for Peak current Vs square root of scan rate. I have found it non linear. Why it is? Is it showing that the ongoing electrochemical reaction process is irreversible? When i have plotted the Peak current Vs scan rate, it is slightly linear. Why the change occurs? I need the exact reason behind this. Kindly suggest (with reference if possible).
I have attached the file for reference.
Thanks
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Agree with the above. If there is a linear dependence of peak current with sweep rate then it usually means an adsorbed species is involved. This is a good recent paper in the Journal of Chemical Education.
Watch your units on your graph, sweep rate is mV / s.
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Hello scholars,
Recently, I have been working on AOR. I have obtained interesting results, however, the result is a bit disturbing as it comes with noise in the curve. Below is a picture of such. I have read up some of the suggestions presented to similar circumstances in RDE measurement for ORR. I have not been able to overcome the challenge. For my circumstance, am working using Pt wire/coil as counter, Ag/AgCl as reference and GC as working electrode in 1M NaOH.
NB:
1. No leakage in RE as I have checked a couple of times and I have about 3 of these REs at my disposal, so I can interchange and check if there is any leakage (issues) with either of them. The aligator clips look good and have also interchanged with spare.
2. No mechanical vibration / interference as the lab is set up in the basement of the building and the experimental setup is on a standardized experimental bench. In addition, am not working on a hanging electrode.
3. From the picture, I have worked between -0.7eV and 0.6eV. However, I do work also in the range of 0eV and 1.4eV
4. The experimental solution has been prepared hours prior to the the measurement, and given enough time to acclimatize to the room temperature. As regard to the GC electrodes, I follow the standard rules. Polishing via the "8" pattern using 0.05um polishing alumina once in 5 days on the polish and DIW for rinsing.
Do you have any other suggestion?
Thank you
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Try to increase the sample interval and sensitivity to 0.005 it might help (I remember something regarding the ratio between scan rate and sample interval)
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is there any publish research show the salivary biomarkers concentration for breast cancer?
iam trying to design a salivary biosensor for early detection of breast cancer. recommend me some articles please
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I have an electrolyte membrane (~ 200 μm thick) containing Nafion and ionic liquid (EMIM-BF4 ). I want to check ions transfer inside the membrane when it is subjected to an electric field. I'm wondering if there is any way for real-time monitoring of the cations and anions movements toward corresponded electrodes.
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It's possible that an electrochemical impedance spectroscopy analysis (EIS) could provide you useful information (specially a four electrode experiment)
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Biosensors and Bioelectronics 25 (2010) 1301–1306 ; they also calculated the same parameter but i am confused with the units that they put into the equation!
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Hi Md. Musfiqur Rahman
In general, the concentration of electro active sites (Surface coverage) calculated by following equation
Γ = Q/nFA
Where Q is the charge (refer cv data), n is the number of electrons , F is the Faraday constant and A is working electrode surface area.
Hope these information's are helpful to you
Best wishes
M. Thiruppathi
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If Beethoven was alive today, which technology would help him to hide deafness?
According to statistics, 80% of those who could benefit from a hearing-aid choose not to use one. One of the most important reasons for that, is the social stigma associated with common misconceptions about wearing hearing aids. How can bioMEMS help to fabricate miniaturized hearing aids without compromising performance?
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I agree with Ozberk Ozturk.
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My work is on biomolecule sensing by potentiometry (CA, CV, etc.). Concentration of each individual biomolecule can be easily monitored by variation in current or voltage, but how to check selectivity i.e. which specific biomolecule is present in a mixture of different biomolecules. In case of heavy metals the redox potential difference is high which is not the case in biomolecules. So please suggest me how to sense or discriminate specific biomolecule (ex. amino acid).
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Hi Ishu Singhal ,
I have few suggestions for you.
You must do (know) the following:
1. What is the charge of your molecule (analyte) at the working pH? (you can get this from the pka of the molecule).
2. What are the charges of the other interfering molecules.(you can get this from the pka of the molecules).
3. Does your molecule have specific interactions such as pi-pi with your electrode material or thin film.
4. Are there any materials in the literature that showing selectivity towards your analyte.
5. Design an electrode material based the factors above.
6. Use Differential pulse voltammetry technique to know the selectivity of the platform towards your molecule.
7. Do some control experiments such as blank, molecule of interest, and the interfering molecule one by one. Compare their responses and come to a conclusion.
I hope that these steps will surely help you.
Have a great time...
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Hi,
I'm working on an electrochemical biosensor integrated into a microfluidic system. The working and counter electrodes are gold, and the reference electrode is Ag/AgCl.
I'm planning to either immobilize thiolized nucleic acids on the bare gold WE surface, or use 11-mercaptoundecanoic acid SAM method to functionalize the WE surface for EDC-NHS coupling to proteins.
Due to the way the microfluidic device is fabricated, I may need to immobilize the biorecognition probes within closed channels (post-bonding). I also want to functionalize only the WE, while keeping the CE/RE uncoated.
Can anyone guide me on how this can be done?
Thank you,
Afiq
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Hi Afiq,
You can make electrochemical desorption of 11-mercaptoundecanoic from the CE. The electrochemical desorption can be done by sweeping the potential between 0 to -1.4V in 0.1M KOH solution. You should optimize the potential window.
Good luck
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We have transgenic E. coli and we need to isolated and purified the specific protein. We used membrane filtres, however it didn't work as we expected. We don't want to use chromatography for purification, first. I just looking for some other alternatives.
Thank you for response.
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I am not sure if I understand your question correctly. You objective is to isolate & purify the target protein. Filter/Chromatography is not what you want, however usually biosensor is good at isolation rather than purification. I do see some "catch and release" system for biomarker released from live cells.
For example:
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1. The system consists of a protein complexed on thiolated SAM (mercaptohexadecanoic acid) after activation with EDC (carbodiimide). The immobilization is done on a gold coated slide which is the working electrode (WE). The electrode area ~ 1 cm-2.
2. Reference electrode is Ag/AgCl wire and counter electrode (CE) is a Platinum wire.
The protein is further complexed with a DNA aptamer.
Then successive EIS scans were run on the system with 1mm Ferro/Ferri redox couple in 1 X PBS.
However, the system is not stabilizing over more than 2 hours. The Rct and Cdl keep on increasing.
EIS parameters : Freq range : 10 KHz to 0.1 Hz ; AC voltage 5 mV ; No DC voltage
Relevant figures are attached.
What could be the reason behind the drift of the system ? What could be a possible solution. I can wait for longer times but would like to fix this problem.
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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.
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Hi Everyone, Am looking for a Methods / Protocols for Electrochemical Biosensors, can some one shed the light on these aspects,since am new to Electrochemical biosensors , my aim is to build a Electrochemical Biosensor to detect microbial pathogens.
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Thanks Irene
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I am facing a big problem regarding fitting and modeling my impedance data. I am using a non-Faradaic system (only filtered and degassed 10 mM PBS solution) for the EIS measurement of gold disk electrode (1.6 mm diameter) modified with SAM, linker, and proteins. I am using the modified Randles' circuit model to fit the impedance curve and get the Rct to plot the dose response curve of my sensor but some paper say that Rct for the non-Faradaic system are too big and almost neglected, usually, the capacitance is used to interpret data. I couldn't find relevant papers using this kind of system and still not sure whether my circuit model is correct for my system or not. Could anyone be able to explain this situation better???
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Dear Roshan,
Thank you for running the CV's as Hunter suggested. It is something that should become second nature. The oxidation/reduction of gold is shifted to more negative voltages as a function of increasing pH, and in the presence of chloride it is further decreased (as compared to sulfate) the standard potential being about 1V vs .SHE. So it is likely that some redox activity comes from Au. I assume you are using a solid gold elctrode, so if you scan to more positive potentials you should be able to resolve the full oxidation peaks (if you use thin film electrodes do not try this, in the presence of chloride). Another feature of the CV's is the appearance of a relatively well defined reduction peak at about -0.3 V, followed by a constant current this could be related to Au reduction and limiting proton concentration. From the CV of the modified electrode I see no evidence for any reductive or oxidative desorption of the thiol monolayer (but possibly these are steady state voltammograms). So either this does not occur or the modification is imperfect.
Anyhow, the potential you are using now seems to be fine. So a next step could be the characterization of your monolayer. When you read literature the procedure of fabricating a SAM on gold, seems very easy and sometimes it is just assumed that it is perfect. Of course it is almost never the case. So if possible you could characterize your SAM using a redox couple like Fe(CN)63-/4- (1mM) or Co(phen or bpy)32+/3+ (e.g 0.1 mM). Scan limits about -0.1 to 0.4 V vs. AgCl, and EIS at equilibrium. Possibly you might consider to work at low ionic strength (+- 10 mM), for this. And you can then do the same with the attached proteins. If you find the modification to be unstable you can consider using mixed (mono)layers, in the past I found mercaptopropyl-methyldimethoxysilane very useful for blocking and stabilizing purposes. I used this in conjunction with sodiumborohydride reduced antibodies (without further purification), and it worked fine. For cleaning gold sweeping in 1M H2SO4 is very effective and allows you to determine the surface area of the gold electrode, if this is not possible methanol/HClO4 cleaning was also satisfactory.
Hope this helps,
Kind regards, Mike
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I haven't been able to find a "field applicable" phosphate sensor in the market yet. Seems there are many unsolved challenges such as longevity, signal drift, selectivity, etc for this sensor in field. Do you know of any commercially developed field applicable phosphate sensors? Or the reasons why this type of sensor is hard to develop for field applications? Thanks
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I also don't know of any commercial sources for this electrode.  One reason that it is so difficult to develop an electrode for phosphate is its charge.  While, for the most part, monovalent and divalent ions are relatively easier for preparing electrodes, trivalent ions are prone to many difficulties, not the least of which is selectivity.  In addition, as you may remember from the Nernst equation, the response slope of a trivalent-ion electrode would be less than 20mV/pPO43-.   Thus, the sensitivity of such an electrode would be lower than that of lower valency electrodes.
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I just bought the QCM cell from CH instrument and I do not know how to use it in parallel with Cyclic Voltammetry. I am using CV for metal-decorated graphene to enhance the electrode sensitivity toward selective biological metabolite.
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actually EQCm is Cv where you monitor the change of current and the mass of the electrode as well.
so you scan between potentials E1-E2-E3-E4 (depending on your software and you register the values of delta M and current density.
if you have any inquiries then be free to ask.
of course you have to deposit your working electrode (metal) either Cu, Fe, Al etc. because the crystals you buy are mainly Au surface (prefered on Ni).
but some working electrodes are available commercially too (expensive).
good luck.
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Dear researcher
i am working on electrochemical detection of DNA and proteins after adsorption of Fc-DNA probe or Fc-aptamers on reduced graphene oxide modified electrodes. But, the sqaure wave voltammetry signal is not stable with time. always there is 5 to 10% decrease in the swv signal which cannot be used for further measurement. does anyone saw this issue before and how can we figure out it. many publications have been published in this field bit none talk about the stability of the adsorbed DNA containing redox tag.
regards
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Dear,
Although my main experience is with gold-thiol chemistry, the problem seems very similar. After modification of the electrode surface there will always be loss due to non specific adsorption. This may be due to weak interactions or crowding of the electrode surface. Depending on your modification method, I would consider a loss of 5-10% reasonable. Normally I used to include "desorption" step in the modification method. This is essentially an additional incubation which mimics the conditions of the test (but without the analyte).  Normally losses should become smaller after this. To distinguish between stability of Fc or the desorption of DNA probe, you could  use a unmodified DNA probe  and use  for example Co(phen) to characterize the electrode. If the loss is similar under these conditions then the problem originates in the probe adsorption not the ferrocene labeling or Fc stability. Hope this helps.
Kind regards,                          Mike
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I've been running EIS on gold chips with the surface modified with thiol SAM and antibody fragments etc.
Some of my EIS runs result in impedance of ~5000-1000 ohms, but then others shoot up to ~30,000-50,000 ohms. Despite all being treated and produced in the same way. My supervisor also made chips himself and ran them with the same issue.
Any ideas what might cause such a dramatic increase??
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Does the increment caused by a particular antibody or other SAM coating? or it randomly happens sometimes with any surface modification? the answer could be different. I believe in the both previous answers provided here, and in addition to their suggestion, I suggest you fill out the unattached area of the electrode by some thiolated conventional materials such as MCH or in the case of antibody use the BSA. to sum up, my guess is the non-oriented assembling of antibodies on the electrode surface. Maybe they attached through their charged side-chains onto the gold surface and not with their thiolated end. Maybe!
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Currently I work with SPGE (screen printed gold electrode) but I still confuse to figure out what is the different between this electrode with Interdigitated array electrode (IDA) since I want to know whether I can work at the same condition between these two. 
Thank you.
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Dear Daniel Martín-Yerga
Thank you so much for the answer. It helped me figure out what should I do with my works.
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I am working on electrochemical non-enzymatic glucose sensor. I don't know how to  measure glucose amount present in the blood and blood serum. How I compare my result with the standard analytical kit result ? Is there any method to detect it and Weather I have to do IT measurement or any measurement? Please help me. 
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When we develop a new method first we need to know the critical point during chemical reaction to avoid interference.
Them we use the sensor in standard well known analyte (in this case pure glucose from oficial company)
The last step is to compare and apply in real sample (serum in this case)
I hope Prosper Kanyong can give you good advise.
Regards
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What are the arguments of making a distinction between the CE and RE if I am measuring non-faradaic processes (i.e. binding events to a surface modified WE)? I understand the arguments for faradaic measurements, that the CE allows current to pass while to ensure the RE can maintain a constant potential. However, if no current is flowing across my electrode interface(s), why would a 3-electrode benefit?
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Three-electrode configuration allows you to better control what is happening on working electrode. e.g. apply a DC bias to polarize the working electrode for certain purpose. Also,  you are able to focus on the change on working electrode and neglect the effects from counter electrode if you are only interested in working electrode.
What do you mean by " no current is flowing across my electrode interface"?
Dan
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In electrochemical cells where reference electrodes are thin films and are not in contact with standard saturated solutions, how reliable would the readings be in terms of having a stable reference potential?
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As long as the concentration [Cl-] does not change, the reference potential should be stable and your measurement should be fine if you are interested in continuous measurement. The effect on you measurement also depends on the potential you are working on, say if your over-potential is 200mv, 10mv shift of reference potential won't hurt too much.   Probably you will find this paper useful: Fully integrated wearable sensor arrays for multiplexed in situ perspiration analysis.
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Hi!, I´m trying to build a low-cost potetiostat using Arduino Uno platform to perform cyclic voltammetry, but I don´t know how to  modify the original sketch for other techniques like anodic stripping voltammetry or linear sweep voltammetry.
Do I have to use a different microcontroller or use more than one for this goal?
Thanks!
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Also, let us propose to add a buffer amplifier[1] before the analog signal input(s) of  the Arduino. Another (better) option might be a differential amplifier[2,3]. It could be a nice project for scholar studies.
1. see Figure 3. An op-amp–based unity gain buffer amplifier  https://en.wikipedia.org/wiki/Buffer_amplifier
2. see Figure 5: Op-amp differential amplifier  https://en.wikipedia.org/wiki/Differential_amplifier
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I am trying to develop electrochemical biosensor using screen printed electrode. The sensor gives good response in blank solutions. However, in presence of proteins, such as albumin, the signal completely disappears. I want to understand if there is a way to prevent protein adsorption, which I believe is distorting the signal.
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By mixing your modifier with charged polymers (such as Nafion and chitosan) you can increase the chances of electrostatic repulsion. Nafion is negatively charged polymer (sulfonate); it will electrostatically repel other anions or negatively charged species , like ascorbate, urate. Moreover,chitosan is a  positively charged biopolymer that can be used for your work.
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In an electrochemical reaction O+ne=R, in the anodic direction (R->O+ne), above a certain level of applied potential, system hits the mass transport limit. Then, bulk concentration of O won't change the current density. How can we experimentally test this to make sure that variations in O does not affect current density? I know that sampled current voltammetry gives some clue, but what if the system operates near the limit and even small variations in signal matter?
Thanks
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If you use a cyclic voltammetry, in a cyclic voltammetry experiment, the solution is kept unstirred; in this situation, mass transport can occur only by diffusion due to concentration gradients created around the electrode surface.
If it is a reversible system,
I will use this eq.
Randles–Sevcik equation:
Basically reversible (diffusion-controlled) and irreversible (charge-transfer controlled) kinetics of electrode process:
Peak Current (i) will be proportional to square root of scan-rate.
i = v1/2  , simplified form of RS eq.
Hope it helps!
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I have got Rct value of bare GCE of 80 kohm cm2. Please help me to explain why Rct value is high for bare GCE?
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Cleaning is critical step. Some day we should spend time to do it. After polishing remove all AL resíduos with ddwater
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Brief system description :
Gold coated glass slide is the working electrode. Used an Al foil clamped to the Au coated glass for connecting to the working electrode of the potentiostat. A part of the gold surface is used for holding a 1 ml drop of 1mM ferro/ferri cyanide in 1X PBS buffer.
Ag/AgCl is used as a reference electrode and Platinum wire is used as counter electrode.
Experiment:
1) Firstly EIS with 5 mV AC perturbation was done on the Au coated glass slide. (bare)
2) Then the electrolyte from the surface was removed and 300 micro liter of 3 mM mercaptohexanol was immobilized for 1 hour. Then it was washed in dd water, dried in air and again the ferro/ferricyanide in 1X PBS was put in. EIS is run with the same parameters as step 1.
3) The SAM layer immobilized Au coated glass slide was heated in acetone solution at 70oC, washed in water and step 1 was repeated.
According to my hypothesis SAM layer will increases the impedance and thus increase the "charge transfer " resistance. However, I am observing the reverse phenomenon for my system.
The plot is attached.
What could be the cause ?
Reference for hypothesis :
Shinn-Jyh Ding, Bin-Wha Chang, Ching-Chou Wu, Min-Feng Lai, Hsien-Chang Chang, Impedance spectral studies of self-assembly of alkanethiols with different chain lengths using different immobilization strategies on Au electrodes, Analytica Chimica Acta, Volume 554, Issues 1–2, 4 December 2005, Pages 43-51, ISSN 0003-2670, http://dx.doi.org/10.1016/j.aca.2005.08.046.
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Yes, a little bit. The larger the surface, the harder it is to passivate it totally. Can you reduce the electrode size significantly and see if you still have the same problem?
About your electrolyte, yes ferro,ferricyanide is good. The contration and buffer are good as well. I'm assuming 1X means 100 mM. Keep in mind that the current you get will increase with buffer salinity. In your case, with a large electrode, the current might be large. Do you have an order of magnitude?
About your thiol. It has OH groups, they are charged and can be protonated but at pH=7.5, you should be mostly ok. 
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I want to test the antioxidant capacity of plant-derived alkaloids (conjugated with gold naoparticles by green synthesis) on electrochemical biosensor using Xanthine Oxidase-Superoxide Dismutase enzyme system. What could be the best strategy for screen printed electrode?
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Dear Muhammad.,
In general carbon porous materials (as graphite) are really suitable for immobilization of enzymes. On the other hand, on vitreous carbon materials it is more difficult to achiever big loadings of active enzyme on the electrodes.
So the first question you should answer is, what type of carbon material is your screen-printed electrodes made of?
The second question you have to face is if your enzymes system is going to transfer the electrons directly with the electrode or if you need a mediator. In case you need a mediator, would you immobilize it in the electrode or would you have it in solution?
If your carbon is porous then simple adsorption of the enzyme system on the electrode would be feasible. If your carbon electrode is vitreous you would need some type of modification on the electrode or incorporate the enzyme with polymers or gels on the electrode.
I hope this helps.
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When I used oxygen plasma treatment for sticking PDMS to glass substrate which has Au-Cr electrodes, the electrodes became damage. How can I solve this problem?
In the attached photo, the device which was used for making plasma is shown.
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You might try reducing your RF power. What equipment are you using for plasma treatment? Time may also be reduced. Some people have reported treatments as long as 60S in Harrick palsma cleaners for instance, but we find that 20S works best for us.
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I tried Nafion, the binder, but it fell off.
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David; Nafion coating/film is used to improve the selectivetity of cations over ions.
But I do not know how to do the coating myself.
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Hello,
I have a device resembling the one in the attached image. I want to measure the current across the two Ag/AgCl electrodes separated by the membrane at different voltages (e.g. +200 mV , -200 mV) etc. 
The membrane is Au coated and is nanoporous in nature. It will be immobilized with organic molecules in later experiments.
I have a Gamry Potentiostat in my lab which has a program called set voltage. Can I use that for performing the experiment ?
I am unsure about the technique as I have little or less knowledge in electrochemistry.
Thanks.
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Never use Ag/AgCl electrode for this experiment (will be destroyed after the first run). Consider 2 Pt electrodes (large area). Use AC current to measure the ionic conductivity.
The membrane must be the  limiting factor for your ionic current (not the solution).
OPTION A:
Use AC current to measure the ionic conductivity. Take the signal from a  conductometer (25…1000 Hz). No electrochemistry (electrolysis) is involved in such an experiment.
OPTION A2 (most recommended)
Adapt  the cell of a conductometer for these measurement (with/without  membrane).
OPTION B:
Use the EIS facility of your potentiostat if available. Connect one Pt to WORK and REF and AUX together to the other Pt. Choose E= ocp, AC signal 10…25 mV, f= 100 kHz….1 Hz. In the Nyquist Plot read R1 (what is called solution resistance) for further comparison. Measure with and without he membrane to see if it is working.
OPTION C (less recommended)
Use cyclic voltammetry (a solution of Fe2+/Fe3+) in a 3 electrode cell with/without membrane).
Think to other models for your experiment. The electronic current in the external circuit is equal to the ionic current in solution (compare results with/without membrane). Maybe the aim of your experiment is not to measure currents (depending on voltages), but measuring directly the conductivity of your membrane (not depending on applied voltages). Think also to osmosys as a starting idea.
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Hai,
I am performing sandwich ELISA to detect biomarkers (as antigens).  
The cyclic voltammetry(CV) of the the SPCE is taken to analyze the analyte (biomarker).  
The primary antibody is immobilized onto the working electrode.
What are best way to chemically pre-treat carbon electrodes before using EDC-NHS.
What chemicals are suitable for this pretreatment?
I appreciate your time. Please reply.
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In order to use EDC-NHS chemistry as activator / coupling agent, your matrix needs to have accessible carboxyl groups.
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Hello, I' m searching for details about graphene based biosensors for the detection of glucose. Can anyone tell me about the electronic properties related to the application and if there are any factors that limit the deployment of graphene in glucose biosensing? My research is referring to non-composite graphene biosensors. The most research papers I studied refer to graphene oxide or praphene composites.
Thank you in advance
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I read papers on reducing graphene oxide using glucose, so i think you can use graphene oxide instead of graphene.  With graphene you should know the effect of glucose on reduced graphene.