TITLE: Automated H&E Staining and Coverslipping (Leica)
AUTHORS: Linda Thomas, Angie Brown, Kerry Wiles

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This procedure establishes a consistent process for preparing H&E slides from FFPE tissue samples using the automated H&E Leica Stainer and coverslip instruments. This procedure is performed by trained histology laboratory personnel.</div></div>

[STEPS]
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Turn on the Automated Robotic Cover slipper and check levels for mounting media and coverslip. After initializing, retrieve the white brush clip from the Xylene well in the drawer on the front and clip it in place.
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Take the needle out of the small xylene bottle and press the ‘Prime Needle’  button while holding the needle over something to catch the xylene and glue that will come out. An empty coverslip container is kept for this purpose.
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Move the needle over its holder and push it down into place, aligning the grooves on the side of the needle mechanism with the metal pegs.
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Make sure that the cover-slipper racks are in place to catch the slides.
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Load 5-6 blank slides in a Stainer rack. All slides should be facing the ‘Up’ label. Load rack into the Xylene well in the drawer and press start. Remove rack when coverslipping is complete and discard the blank slides.
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Open the lid on the Leica Stainer and remove the metal reagent covers. Turn the machine on.
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The screen should show a representation of the reagent wells. The only program we use to stain is the H&E program, and it should already be set to run. Do not change any of the settings on screen.
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Place a red clip on the front of the rack (The side labeled Up). The red clips must be used with the H&E program. The clip color corresponds to the program being run. The wrong clip color will disable and prevent the Stainer from running the program.
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Open the drawer on the front of the machine and place the racks inside. The red clips should be facing away from the machine.
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The slides will move from the Stainer to the coverslipper automatically. The order the slides are dipped into the reagents and the amount of time they spend in each reagent is set in the program. Once the process has begun the user does not need to do anything to assist the process unless the coverslipper rack fills up, in which case an alarm will sound.
Staining Protocol   ABC1STEPSTATIONTIME211-Xylene3:00322-Xylene2:30433-Xylene1:30544-100% Alcohol1:00655-100% Alcohol2:00766-95% Alcohol 1:00877-95% Alcohol1:009812-Wash0:3010922-Hematoxylin 3:00111010-Wash1:30121121-Clarifier1:0013129-Wash 1:00141320-Bluing Reagent1:0015148-Wash1:00161519-95% Alcohol1:00171618-Eosin1:00181717-95% Alcohol1:00191816-100% Alcohol2:00201915-100% Alcohol2:00212014-Xylene 2:00222113-Xylene1:002322Coverslipper
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When the coverslipper sounds an alarm, the Stainer will also sound an alarm to indicate that coverslipper needs attention. Turn off the alarm on the Stainer by hitting the back arrow to get rid of the message that says the coverslipper needs attention.
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Hit the alarm button on the coverslipper to turn it off.
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Carefully remove the full racks from the coverslipper and replace them with empty ones.
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Once the new racks have been placed, hit ‘Start’ to continue the process.
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When the machines are done, turn off the Stainer and place the metal well covers over the reagents to keep them from evaporating. When the Stainer is turned off, sometimes the cover-slipped alarm will sound. If this happens, turn off the alarm.
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Remove the emptied Stainer racks from the machine.
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Go to the coverslipper, place the needle in the small bottle of xylene kept inside the machine.
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Take the racks out of the coverslipper and set them aside to the side to dry.
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Remove the white brush clip and place it in the xylene well. Place the cover over the xylene well before closing the drawer.
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Turn the coverslipper off.