TITLE: Cyclic Voltammetry analysis of laser-inscribed graphene electrodes: Ruthenium(III) chloride trihydrate redox couple
AUTHORS: Nick Cavallaro, Carmen Gomes, Eric S McLamore

[DESCRIPTION]
This protocol describes use of the cyclic voltammetry (CV) method for electrochemical analysis of laser-inscribed graphene (LIG) electrodes using hexaammine-ruthenium(II) chloride (Hills et al; Oliveira et al). The protocol requires approximately 1 hour (excluding electrode fabrication).

[BEFORE_START]
Always wear proper PPE during experiment (gloves, eyewear, lab coat, proper clothing)

[GUIDELINES]
The protocol is based on numerous published manuscripts that use similar methods with the haexamine-ruthenium redox couple, but is not a comprehensive guide. Where appropriate, alternative redox couples or electrolytes are noted.


[STEPS]
SECTION: Preparation
1. Prepare LIG electrodes 
Use protocol for LIG fabrication to prepare a batch of electrodes (link here).
 

Expected results from step 1:
Fig 1 shows photographs of the instrument and the expected results from a batch fabrication of six replicate electrodes based on the LIG fabrication protocol here 

 

 

For quality control process to prepare large batches of LIG electrodes, see Qian et al (2024) (link). As an example, Fig 2 shows a heat map from the auotmated batch clustering tool. 


 

SECTION: Preparation
2. Prepare solution
Gather a sealed glass storage bottle or Falcon tube with at least 50 mL of capacity 
Label as 4mM [Ru(NH3)6]Cl2 + 100mM KNO3
In a separate weigh boat/paper, prepare the following:
                1-Weigh 54.6 mg hexamine-ruthenium chloride on a scale.
                2-Weigh 505.5 mg potassium nitrate on a scale.
Add the potassium nitrate to the labeled glass bottle. 
Fill with 25 mL of DI and stir/agitate well until solution is dissolved
Add the hexamine-ruthenium chloride. 
Fill with an additional 25 mL of DI (50mL total volume) and stir/agitate well until solution is dissolved
Transfer 20 mL of the solution to the electrochemical cell (20 mL cell shown in Tang et al protocol)

 

SECTION: Electrochemical testing
3. Connect electrodes
Assemble electrochemical cell (Benchtop Palmsens4 system):

Single electrode system
Connect the single LIG electrode with the potentiostat via the bonding pads using alligator clips
Ensure the electrodes are not touching, and the distance between the auxiliary and working electrodes is consistent. 
Tang et al protocol shows the correct assembly of the electrochemical cell for a single LIG "dip-style" electrode, where a Ag/AgCl reference electrode and Pt wire are used as reference and counter electrode, respectively

 

SECTION: Electrochemical testing
4. Conduct scan (PalmSens4 benchtop system)
 

Turn on Palmsens hardware and the computer. 
Open the PalmSens software (MultiTrace4 software).
Users may operate 3 channels simultaneously or individually
Select Cyclic Voltammetry as the Technique. 
After selecting the mode, set up all parameters. For example, see settings below (see MultiTrace manual for details; link here)

 

Expected results from PalmSens4 benchtop potentiostat
Fig 3 shows the channels connections to the PalmSens, mode selection panel, and operation panel.
When the experiment finishes, click export to excel icon to export data.
When exporting data, make sure all channels' data are exported.

 

SECTION: Clean up
5. Clean workspace (Timing: 5 min)
Ensure chemicals are stored or discarded correctly, and turn off Palmsens and the computer.
If using external reference and auxiliary electrodes, clean them up with DI water and store them in the appropriate place. Reference electrodes should be stored with a 3M KCl solution.
Wipe the working desk with 75 % alcohol. 
Store electrodes when not in use.