Let people know what your project can do specifically. Provide context and add a link to any reference visitors might
be unfamiliar with (for example your team wiki). A list of Features or a Background subsection can also be added here.
If there are alternatives to your project, this is a good place to list differentiating factors.
A PCB in the shape of a microscope slide harbouring 5 bridgeable gold electrode. It can be used with the [AC Dispatcher (ACD)](https://2022.igem.wiki/paris-bettencourt/ACDispatcher) and the [Electro Planner](https://2022.igem.wiki/paris-bettencourt/Electro_planner) to expose cells to electrical signal in solid medium.
## Installation
Within a particular ecosystem, there may be a common way of installing things, such as using Yarn, NuGet, or Homebrew.
However, consider the possibility that whoever is reading your README is a novice and would like more guidance. Listing
specific steps helps remove ambiguity and gets people to using your project as quickly as possible. If it only runs in a
specific context like a particular programming language version or operating system or has dependencies that have to be
installed manually, also add a Requirements subsection.
### Quick Links
## Usage
Use examples liberally, and show the expected output if you can. It's helpful to have inline the smallest example of
usage that you can demonstrate, while providing links to more sophisticated examples if they are too long to reasonably
include in the README.
**Used for:**
## Contributing
State if you are open to contributions and what your requirements are for accepting them.
Inputs
Screening
For people who want to make changes to your project, it's helpful to have some documentation on how to get started.
Perhaps there is a script that they should run or some environment variables that they need to set. Make these steps
explicit. These instructions could also be useful to your future self.
The Electo-Micro-Slide is a microscope slide of a new kid. This Printed Circuit Board, taking the exact dimension of a standard microscope slide, therefore fitting in a microscope slide holder present in most fluorescent microscope. The Electro-Micro-Slide receive electrical signals from the AC Dispatcher (ACD) and through 5 individual pins connected to physically segregated pairs of non connected gold electrodes. Connection between these electrode is done though depositing an agar pad across the pair of electrodes.
Holes between the each pair of electrodes allow for light to pass in both direction and for illumination through trans and fluorescent lighting of the studied sample. We used the Electro Planner to shock the program the experiment
The device is comprised of a 6 pin IDC connector for easy connection to the ADC through a flat ribbon cable, rendering manipulation in often crowded microscope chamber easy and tidy.
The device is largely inspired by the work showcased in \[1\]. However, as the device proposed by Stratford et. Al is manufactured through Vapor deposition, fabrication of their tool is impossible to many labs. Wit h the specific vision to enable these kind of experiments in labs beyond the fields of life science (such as in Robotics), we redesigned their device so that it could simply be ordered already manufactured and assembled, by the click of a button and for a very accessible price (less than 1$ per piece).
Experiments done with it:
-------------------------
We orignally developed this device to invcestigte the possiblity of controling the expression of the pNasA promoter in _B. subtilis_ though hyperpolarisation induced starvation of glutamine \[2,3\]. However, because of time constrain, we didn’t arrived to the point where we managed to assess the viability of this strategy.
We would like to point out nonetheless, that we have submited this idea to [Muhernio Asally](https://scholar.google.com/citations?user=vKC8SbkAAAAJ&hl=fr), author and director of the lab that produced the works affored mentioned, trhough a very graciously granted interview. His view on it is that it sounded feasible and, if we will get more opportunity in the future, we will attenpt this experiment.
In our work, we have used the ACD in conjunction with the Electro Planer, the HTEA and the Micro-Electro-Slide. Many other applications remain highly possible.
Materials and Methods
---------------------
We electroshocked bacteria stained with 10 uM Thioflavin T (a positively charged cationic dye reporting membrane potential) at frame 4.
Images were obtained though fluorescent microscopy (exposure for GFP Excitation and emission) on the Nikon XXXX at 60X Magnification
Cells were exposed to electrical signals generated by the MSH-2300q and controled by the [Electro Planner](https://2022.igem.wiki/paris-bettencourt/Electro_planner). These signals were gated and dispatched by the to [AC Dispatcher (ACD)](https://2022.igem.wiki/paris-bettencourt/ACDispatcher) the [Electro-Micro-Slide](https://2022.igem.wiki/paris-bettencourt/Electro_Micro_Slide)
Cells were exposed to signals of varying amplitude (from 0.5 volts to 6 volts) at 100HZ and for 2.5seconds.
Results
-------
We can observe that the shock provoke a slight displacement of the entire field of view. The fluorescence intensity also change as expected though experiments conducted in \[1\]. However, because of the displacement, drawing conclusions from this experiments on the ability to change membrane potential is to be taken slightly
**Overall, we showed that:**
* observing bacteria's on the microscope with the Micro-Electro-Slide was achievable.
* Exposure of bacteria to electrical signals through conjoint used of the Micro-Electro-Slide, the ADC and the Electro-Planner have the desired effect
**_E. coli_ observerd thorugh one of the wells of the Micro-Electro-Slide** 
**Video of _E. coli_ observerd thorugh one of the wells and receiving an electroshock**
References
----------
1\. Stratford, J.P. \*et al.\* (2019) ‘Electrically induced bacterial membrane-potential dynamics correspond to cellular proliferation capacity’, \*Proceedings of the National Academy of Sciences\*, 116(19), pp. 9552–9557. Available at: [https://doi.org/10.1073/pnas.1901788116](https://doi.org/10.1073/pnas.1901788116)
2\. Prindle, A. \*et al.\* (2015) ‘Ion channels enable electrical communication in bacterial communities’, \*Nature\*, 527(7576), pp. 59–63. Available at: [https://doi.org/10.1038/nature15709](https://doi.org/10.1038/nature15709).
3\. Liu, J. \*et al.\* (2015) ‘Metabolic co-dependence gives rise to collective oscillations within biofilms’, \*Nature\*, 523(7562), pp. 550–554. Available at: [https://doi.org/10.1038/nature14660](https://doi.org/10.1038/nature14660)
function reply\_click(clicked\_id) { var button\_count = 3 var index = parseInt(clicked\_id) // alert(index); var sub\_address = "sub\_button" var master\_address = "button" for (let i = 0; i < button\_count; i++) { address = master\_address + (i + 1).toString(); console.log(address) var doi = document.getElementById(address) doi.style.display = "none" address = sub\_address + (i + 1).toString(); console.log(address) var doi = document.getElementById(address) doi.style.display = "none" } // console.log("address") address = master\_address + index.toString(); console.log(address) var doi = document.getElementById(address) doi.style.display = "block" address = sub\_address + index.toString(); var doi = document.getElementById(address) doi.style.display = "block" } var graphs = ; Plotly.plot('chart1', graphs, {});
> If your team does not have any software tool, you can totally ignore this repository. If left unchanged, this
repository will be automatically deleted by the end of the season.
### In Brief
An open source potentiostat we have characterised and adapted to global current shortage of semi conductors and electronic parts.
It can be used both as a sensor for the Output devices ( [Electrical Resistance as a measure of Growth](https://2022.igem.wiki/paris-bettencourt/Electrical_resistance_growth_mesure)) of our [Electro-Genetic Framework](https://2022.igem.wiki/paris-bettencourt/toolkit) and as an actuator for the Input Devices of the toolkit (See [pSoxS Experiments](https://2022.igem.wiki/paris-bettencourt/pSoxS)). We used this device in most experiments involving redox sensing mechanism.
### Quick Links
## Description
Let people know what your project can do specifically. Provide context and add a link to any reference visitors might
be unfamiliar with (for example your team wiki). A list of Features or a Background subsection can also be added here.
If there are alternatives to your project, this is a good place to list differentiating factors.
**Used for:**
## Installation
Within a particular ecosystem, there may be a common way of installing things, such as using Yarn, NuGet, or Homebrew.
However, consider the possibility that whoever is reading your README is a novice and would like more guidance. Listing
specific steps helps remove ambiguity and gets people to using your project as quickly as possible. If it only runs in a
specific context like a particular programming language version or operating system or has dependencies that have to be
installed manually, also add a Requirements subsection.
Inputs
Screening
Output
## Usage
Use examples liberally, and show the expected output if you can. It's helpful to have inline the smallest example of
usage that you can demonstrate, while providing links to more sophisticated examples if they are too long to reasonably
Potentiostats are standard equipment in electro-chemistry laboratories, but the price generally ranges from 1,000 to 10,000 €. As Electro-Genetics is a nascent field, rare are the biology labs equipped with such devices and their cost is a barrier to entry for most iGEM projects. The IO Rodeo potentiostat is a well-characterised and published in peer-reviewed scientific journals\[1-5\] open-source project comprised of a main Printed Circuit Board (PCB) harbouring the Operational Amplifiers (OP Amps) central to potentiostat operation and extension capabilities allowing for multiplexed measurements of multiple electrodes.
If the assembled devices could (in regular times) be bought from the website for 200$, thus incredibly lowering the barrier to entry to electro-chemistry, the product was out of stock during the period of iGEM 2022 because of an [international shortage of microchips](https://en.wikipedia.org/wiki/2020%E2%80%93present_global_chip_shortage#:~:text=The%202020%E2%80%93present%20global%20chip,affecting%20more%20than%20169%20industries)
After inspection of the PCB Design files, we understood that the missing components were the Quad SPST CMOS Analogue Switches DG411DY-T1-E3 and the Drift Voltage Reference REF3330AIDBZR. We replaced both of them with available Though hole parts. The new PCB (Available on our Gitlab) can be ordered for manufacturing and assembly of all surface-mount parts in online PCB Manufacturers. We ordered 5 of them for the price of 45$ thus massively lowering the price of these devices.
We characterised this device through cyclic voltammetry and constant voltage voltammetry experiments and compared our results with commercially available [PalmSense Sensit BT](https://www.palmsens.com/product/sensit-bt/) potentiostat, graciously lended to us by [Prof. Vincent Noel](http://www.chimie.univ-paris-diderot.fr/fr/annuaire/itodys/vincent-noel) of the university Paris Cite.
We used it in:
--------------
### Calibration of Potentiostat
We prepared a common reference redox solution of ferri/ferrocyanide (K3\[Fe(CN)6\]) to calibrate the IO Rodeo potentiostat to the commercial Palm Sens ⟨™⟩ Sensit BT potentiostat. We performed **cyclic voltammetry, a method often used in electrochemistry to investigate redox systems.** It is a technique for qualitative analysis method of reaction rate and mechanism. By reproducing the same results in our open-source potentiostat as the commercial one, we demonstrate that our potentiostat detects the necessary range of redox reactions.
### Electrochemical gene induction
We used this device in the context of the characterisation of the Input parts of our Electro-genetic toolkit to induce gene expression with an electrical potential (See [pSoxS Experiments](https://2022.igem.wiki/paris-bettencourt/pSoxS)) and in the context of the [lysis based Output part](https://2022.igem.wiki/paris-bettencourt/Electrical_resistance_growth_mesure) of our Electro-Genetic Toolkit.
The commercial potentiostat Palm Sens ⟨™⟩ Sensit BT was lent to us by Vincent Noel, professor at the UFR Chimie (Université Paris Cité). IO-Rodeo was designed on a PCB board and ordered. A stock solution of 2 mM Potassium ferrocyanide (K4\[Fe(CN)6\]) in 1 M Potassium nitrate (KNO3) was prepared following the protocol on this link (\[https://www.als-japan.com/1898.html#defaultTab11\](https://www.als-japan.com/1898.html#defaultTab11)). 3 mL of this solution were transferred into a culture tube. DropSens electrodes were immersed in the solution and connected to the potentiostats.
Cyclic voltammetry was performed first with the Palm Sens ⟨™⟩ Sensit BT, and then with the IO-Rodeo potentiostat. The settings of the commercial potentiostat were selected through the provider software PSTrace, whereas the IO-rodeo cyclic-voltammetry parameters were coded in python. The code for the IO-Rodeo cyclic voltammetry and chronoamperometry was imported from the IO-rodeo potentiostat documentation (\[http://stuff.iorodeo.com/docs/potentiostat/examples.html#cyclic-voltammetry\](http://stuff.iorodeo.com/docs/potentiostat/examples.html#cyclic-voltammetry)) and the parameters adjusted.
**Electronic induction of gene expression**
The IO Rodeo potentiostat, connected to the DropSens electrodes, was used in the pSoxS experiment (https://2022.igem.wiki/paris-bettencourt/template\_experiment) involving the redox species pyocyanin. Cyclic voltammetry was performed in a LB solution with 10uM of pyocyanin, to find the reduction peak. The reduction voltage potential was applied by chronoamperometry (also known as constant voltage voltammetry) for 16 hours to reduce pyocyanin, and activate expression of fluorescence in cells.
Results
-------
**Comparison of cyclic voltammetry between the commercial and the IO-Rodeo potentiostats**
The Cyclic voltammetry graph of the commercial potentiostat (left) shows an oxydation peak at 0.1 V and a reduction peak at -0.05V. On the IO-Rodeo we notice a slight shift in the peak values with the oxydation peak at 0.25V and the reduction peak at 0.12V. These values are not absolute and represent a qualitative measure of redox activity.
**Pyocyanin cyclic voltammetry and chronoamperometry**
In the pyocyanin cyclic voltammetry graph we notice a oxydation peak at 0.3V and a reduction peak at -0.5 V. This relative value was used for the chronoamperometry experiment to reduce pyocyanin.
1\. Bullen JC, Dworsky LN, Eikelboom M, Carriere M, Alvarez A, Salaün P (2022) \*\*Low-cost electrochemical detection of arsenic in the groundwater of Guanajuato state, central Mexico using an open-source potentiostat\*\*. \*PLoS ONE\* 17(1): e0262124. \[https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0262124\](https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0262124)
2\. Fatoni A, Widanarto W, Anggraeni MD, Dwiasi DW (2022) \*\*Glucose biosensor based on activated carbon – NiFe2O4 nanoparticles composite modified carbon paste electrode\*\*\*Results in Chemistry\*, Volume 4, 100433 \[https://www.sciencedirect.com/science/article/pii/S2211715622001527\](https://www.sciencedirect.com/science/article/pii/S2211715622001527)
3\. Bogoslowski, S., Geng, F., Gao, Z., Rajabzadeh, A.R., Srinivasan, S. (2021). \*\*Integrated Thinking - A Cross-Disciplinary Project-Based Engineering Education. \*\*\*\*In: Auer, M.E., Centea, D. (eds) Visions and Concepts for Education 4.0. ICBL 2020. Advances in Intelligent Systems and Computing, vol 1314. Springer, Cham. \[https://doi.org/10.1007/978-3-030-67209-6\_28\](https://doi.org/10.1007/978-3-030-67209-6\_28)
4\. Fatoni A, Wijonarko A, Anggraeni MD, Hermawan D, Diastuti H, Zusfahair (2021) \*\*Alginate NiFe2O4 Nanoparticles Cryogel for Electrochemical Glucose Biosensor Development.\*\*\*Gels\*. 2021 Dec 17;7(4):272. \[https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8701366/\](https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8701366/)
5\. Guillem P, Bustos RH, Garzon V, Munoz A, Juez G (2021) \*\*A low-cost electrochemical biosensor platform for C-reactive protein detection.\*\*\*Sensing and Bio-Sensing Research\* 31 (2021) 100402. \[https://doi.org/10.1016/j.sbsr.2021.100402\](https://doi.org/10.1016/j.sbsr.2021.100402)
6\. Cordova-Huaman, A.V., Jauja-Ccana, V.R. and La Rosa-Toro, A. (2021) ‘Low-cost smartphone-controlled potentiostat based on Arduino for teaching electrochemistry fundamentals and applications’, \*Heliyon\*, 7(2), p. e06259. Available at: \[https://doi.org/10.1016/j.heliyon.2021.e06259\](https://doi.org/10.1016/j.heliyon.2021.e06259).
7\. Crespo, J.R. \*et al.\* (no date) ‘Development of a low-cost Arduino-based potentiostat’, p. 21.
8\. Tahernia, M. \*et al.\* (2020) ‘A Disposable, Papertronic Three-Electrode Potentiostat for Monitoring Bacterial Electrochemical Activity’, \*ACS Omega\*, 5(38), pp. 24717–24723. Available at: \[https://doi.org/10.1021/acsomega.0c03299\](https://doi.org/10.1021/acsomega.0c03299)
function reply\_click(clicked\_id) { var button\_count = 3 var index = parseInt(clicked\_id) // alert(index); var sub\_address = "sub\_button" var master\_address = "button" for (let i = 0; i < button\_count; i++) { address = master\_address + (i + 1).toString(); console.log(address) var doi = document.getElementById(address) doi.style.display = "none" address = sub\_address + (i + 1).toString(); console.log(address) var doi = document.getElementById(address) doi.style.display = "none" } // console.log("address") address = master\_address + index.toString(); console.log(address) var doi = document.getElementById(address) doi.style.display = "block" address = sub\_address + index.toString(); var doi = document.getElementById(address) doi.style.display = "block" } var graphs = ; Plotly.plot('chart1', graphs, {});
