Step 0: Loading the sample

If necessary, start ScanImage and BakingTray. The instructions over the following pages are summarized as a checklist.

Turn off the PMTs

Turn on the laser

Hit the "Laser" button in the main view then press "Turn On" on the laser window. After hitting this button the text switches to "Turn Off", as shown above. The power will increase slowly and eventually the modelock indicator will turn from red to green. You can open the shutter now too: there is a second shutter on the path so there is no risk. Select the correct wavelength for this acquisition.

Remove the previous sample (if present)

If it is not already visible, open the "Prepare" GUI by clicking on "Prepare Sample", as shown below.

Enter "0" in the "Z" text entry box of the Prepare GUI to lower the water bath using the Z stage. Hose down the objective with distilled water as shown below, then clean the tip with a piece of folded lens paper. Be gentle: don't push against the tip of the objective with the lens paper. If you do not know how to do this, find an experienced person to show you.

Unscrew the blade holder with the large black thumbscrew and remove it from the vibratome. Take care not to cut yourself. Unscrew the blade retaining screw (shown below). Do not loosen the side screws! Remove the blade and dispose in the sharps bin.

Remove the blade by loosening the top screw. Do NOT loosen the side screws.

Confirm whether the system requires the objective to be raised for the bath to slide out. Then slide out the bath. Dispose of brain slices as appropriate.

Start a new sample

Select Sample -> New from the menu. This will reset the recipe and turn on the resonant scanner (if present). NOTE: you do "New Sample" with the previous sample in place you will have to manually increase the number of sections to slice.

Clamp the new sample into the water bath

Unscrew the clamp holding the slide in and dispose of the slide in the sharps bin. Insert the new slide with the sample into the clamp. Note there is a stop on the slide clamp to keep the slide from protruding over the edge of the platform. The agar block should be placed adjacent to this stop. In the following image, the stop is a round headed screw.

Slide placed into the sample holder pedestal

Push the metal clamp over the slide and tighten it down. Do this carefully: first turn one side until it just starts to tighten and repeat on the other side. Then return to the first screw and tighten until finger tight. Repeat with the other. The slide should now be tight.

Clamp in place and tightened

Place the water bath onto the stage

Gently and carefully place the water bath onto the stage. Push it all the way back into the holder and tighten the retaining screw(s) until just finger tight.

Put in a new blade

Insert a new blade into the holder and tighten until a little beyond finger tight. Thread the blade holder thumbscrew through the holder and screw the holder to the vibratome. Whilst you tighten, push down on the blade holder with two fingers as shown below.

Push down on the blade holder as shown whilst tightening the thumbscrew

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