Make sure you have a surgery room booked

Some sort of table or surgical workbench to transfer rat to when fully anesthetized

Grab isoflurane cart and/or supplies and bring it to the surgery room

Place paper towels in the plastic container

Make sure the tubing is tightly sealed together (if tape is needed add tape)

Make sure all of the tubing is in the correct place and connected

Make sure the valves are parallel with the tubing (the pink and orange tubing)

Use larger thicker tape to secure white tubing down onto the surgery table (where you want it on the workbench)

Snip open a little hole in a glove and place it over the tubing for the rat's mouth as a nose cone for anesthesia during surgical procedures

Check isoflurane level (make sure it is between the minimum and maximum fill lines)

If the isoflurane level is low then unscrew the cap and slowly add in the isoflurane

Set isoflurane regulator on top-adjust dial as appropriate

Use the black wrench to open the oxygen tank

Turn to the right to open

Look at the oxygen regulator and turn the green knob so the silver ball is in the middle of the 1 on the regulator.

Place the rat in the plastic box and close the lid

Adjust isoflurane dial as needed

Start at 4 (if female)

Start at 5 (if male), heavier animals usually need more isoflurane to become anesthetized

Keep rat under for a few minutes or until the rat’s breathing slows down

Take the rat out of the plastic container and place it onto the surgery table

Do a toe pinch using forceps to make sure the animal does not feel any pain

If the rat has a pain reflex or a jerk motion of the limb then place back into the plastic container and continue anesthesia

If the rat has no pain reflex or jerk then continue

Put the tubing that has the glove with the slit over the rat's mouth and nose to maintain anesthesia

Then when the rat is fully anesthetized you can proceed with your surgery

Turn off the isoflurane dial (back to 0)

Take the black wrench to close the oxygen tank

Turn to the left to close

Check to make sure there is no tubing loose/disconnected

In case there is a leak

Or if something is not working

Adjust dial

More or less isoflurane depending on how the rat is doing

Press the green oxygen flush button

This clears out oxygen from the tubing

Remove the paper towels and dispose of them

Clean out the plastic box with paper towels and water only

Return isoflurane machine and supplies to its storage place

Place cage where the dirty cages go to be cleaned