Hi.My name is Resh Gary Dory, and I'm a research assistant in the faf Lab of the Salk Institute for Biological Studies. Today I'll be showing you how to fabricate extracellular suction electrodes using two methods. These electrodes can be used to take electrophysiological recordings of my spinal cords in vitro.
Use an alcohol lamp as a heat source to melt the PE 90 plastic tubing into the desired width. Hold the tubing over the lit flame and remove as it begins to melt, gently pull the ends apart, being careful not to break the middle. This process is shown again.
Remove the tubing as it softens and melts and gently pull the ends apart. Cut the tubing with a sharp razor blade at a point corresponding to the approximate width of the sample to be measured at the thick end of the tubing. Make a cut at a 45 degree angle.
A feral and O-ring must now be installed onto the thick end. Insert this end of the tubing through the hexagonal opening of the feral. Then insert the end of the tubing into the rubber O-ring.
This setup will be screwed into the front of the electrode. Cut a one centimeter length of Zeus small tubing with a sharp razor blade and inspect the ends for a clean cut and an open boar Cut a 10 centimeter piece of the PE 90 tubing using a sharp razor blade. Place a drop of JB weld onto the midpoint of the small tubing using mouth suction.
Draw the small tubing and the weld into the bore of the large tubing and allow this setup to cure overnight. Although it is not shown, a 45 degree cut should be made at the thick end of this tubing, and a feral and o-ring should be installed onto this end. In the same way shown previously.
Disassemble a standard suction electrode unscrew the feral from the front end of the electrode and the BNC connector from the rear. A lower angled side port must now be made on the side of the electrode barrel. Heat up a piece of metal using a bun and burner.
Press the heated metal into the existing hole on the side of the barrel at a low angle. Remove the silver wire from the BNC connector by pulling on the wire where it meets with the connector. Soak the BNC connector in xylene for one hour to remove the wax.
Once the wax is removed, rinse the connector in water and dry with paper towels. Cut a 15 centimeter length of silver wire. Soak the wire in bleach for 15 minutes.
Then rinse and dry with paper towels. Solder the silver wire to the BNC connector. Insert the silver wire into the back end of the suction electrode barrel, and leave enough room for the BNC connector to remain to the side.
Insert the suction tubing into the side port approximately three centimeters. Inject silicone sealant into the back opening of the suction electrode while preventing it from spilling out of the side port by covering it with your finger. Inject sealant approximately two to three centimeters into the electrode barrel.
Once failed, tightly screwed the BNC connector into place, which will push the sealant forward. Let the sealant cure overnight. Screw the feral of the tubing created by either the pulled tip or cut tip method onto the front of the electrode straightening.
The tubing will help ease the silver wire through the electrode is now complete. The recording setup is placed inside a faraday cage to reduce outside interference. Our setup has also been placed on an air table to eliminate mechanical interference.
The electrodes are mounted on magnetic stand micro manipulators and placed near the recording dish. The BNC coaxial cables are connected to a head stage that is connected to an amplifier. The amplifier is wired to a poly view adapter, which is wired to the computer.
This setup is shown closeup. The BNC coaxial cables are connected to a head stage. The head stage is connected to the amplifier.
The amplifier output exits the Faraday cage and is connected to a poly view adapter. The poly view adapter is connected to the computer which has the appropriate recording software installed. The entire setup must be grounded properly.
An external ground is established by connecting the Faraday cage and air table to a ground. An internal ground is established by wiring the dissection dish to the head stage. It is also necessary to ground the doors of the cage to the main body.
With the proper setup data can now be collected. Thank you for watching this video and to help It assist you with your future experiments.
