Dako Autostainer | User Guide Document Number 0003106 Revision E March 2007 For In Vitro Diagnostic Use UL and cUL Dako Colorado, Inc. 4850 Innovation Drive Ft.
Dako Autostainer Information Dako Autostainer Model: _______________________ Serial Number: _______________________ Software Version: _______________________ Computer Configuration: _______________________ CPU: _______________________ Hard Drive: _______________________ Floppy Drive: _______________________ Internal Memory: _______________________ Monitor: _______________________ Dako Autostainer Software Product License IMPORTANT: Please read the license carefully before using this Softwa
Contact Dako Australia Tel. 2 9316 4633 Fax 2 9316 4773 Austria Tel. 0800 0800 7153 Fax 0800 0800 7154 Corporate Headquarters Denmark Tel. 45 44 85 95 00 Fax 44 85 95 95 www.dako.com Distributors in more than 50 countries Belgium Tel. 016 38 72 20 Fax 016 38 72 21 Canada Tel. 905 858 8510 Fax 905 858 8801 Czech Republic Tel. 420 541 423 710 Fax 420 541 423 711 Denmark Corporate Headquarters Tel. 44 85 95 00 Fax 44 85 95 95 Sales Tel. 44 85 97 56 Fax 44 85 84 29 Dako Autostainer | User Guide France Tel.
This page intentionally blank.
Table of Contents Section 1 | Introduction ........................................................................................................ 9 Dako Autostainer Features................................................................................................ 10 Universal System........................................................................................................... 10 Quick Start-up................................................................................................
Adding Slides................................................................................................................. 39 Deleting a Slide.............................................................................................................. 40 The Slide Information Button............................................................................................. 40 Entering Slide Information Through the SLIDE INFORMATION Button ........................
ALL SLIDES Tool .............................................................................................................. 67 Printing a Slide Layout Map .............................................................................................. 67 Section 11 | Reagent Management.................................................................................... 69 Adding New Reagents to Reagent Lists............................................................................
Printing only selected reagents from the Reagent Tracking Report: ........................... 101 Remove Reagent from the Reagent Tracking Log ...................................................... 102 Remove all Reagents from the Reagent Tracking Log................................................ 102 Remove Selected Reagents from the Reagent Tracking Log ..................................... 102 Section 19 | Wash Buffers......................................................................................
Section 1 | Introduction The Dako Autostainer System is an automated slide processing system compatible with currently available reagents for the staining of paraffin-embedded and frozen tissue sections, cytospins, cell smears and fine needle aspirates. This system is designed to automate manual staining methods routinely used in immunohistochemistry and cytochemistry, enabling the transfer of established protocols from the bench to the Autostainer.
Dako Autostainer Features Universal System The Dako Autostainer is a universal system enabling the direct transfer of routine staining protocols from the laboratory bench. Quick Start-up Dako’s Autostainer Support Team installs and tests the system to ensure proper operation. The factory-installed software includes standard reagent protocols and commonly used Dako reagents such as Dako LSAB2, Dako EnVision, and HercepTest™ visualization systems.
Section 2 | System Specifications The Dako Autostainer is a combination of hardware and software integrated with the appropriate reagents to automate immunohistochemical and immunocytochemical staining methods. This section describes the hardware and software specifications and the accessories provided with the system.
Slide Rack Glass Slide Reagent Racks 1 and 2 Reagent Probe Wash Station Sink Y axis mechanism X axis mechanism Z head Assembly 12 is a slide carrier holding 12 microscope slides in a horizontal position. The instrument holds a maximum of 4 racks. These racks are removable and interchangeable for slide loading and unloading. is a standard microscope slide to which specimen(s) is mounted for processing on the instrument. Slide dimensions are 25mm x 75mm x 1mm. hold the reagent vials.
Hardware Specifications Dako Autostainer Dimensions 1.02m W x 0.69m D x 0.61m H 40" W x 27" D x 24" H Weight 140 lb. (63.
Software Specifications Operating System Operating Logic Protocol Logic Idle Rinse Rinse Default Settings Incubation Time Windows® 95/98, or equivalent. Designed to calculate the most time-efficient sequence to complete a programmed staining run. Flexible selection of steps including rinses. Maximum of 35 protocol steps (including rinse steps). Buffer is applied to slides during a staining run to keep specimens wet when no reaction is occurring.
Section 3 | Installation Requirements For installation of the Dako Autostainer the following requirements should be met to ensure that the instrument functions properly. Requirements Surface Size Environment Prior to unpacking the Autostainer, ensure that the area to be used for instrument is a solid, level surface that can safely support the weight of the system (approximately 240 lbs. for instrument plus computer.) Minimum dimensions for the Autostainer work area are 2.00m wide x 0.8m deep x 1.
This page intentionally blank.
Section 4 | Typical Operation A few “off instrument” preparations will make the staining run more time-efficient. Prior to starting a run you should have the following items: • • • • • A list of slide cases to be run with requested protocols. Deparaffinized and rehydrated slides soaking in buffer. This buffer should contain 0.05% Tween 20. A minimum of 5 minutes soaking time is recommended. The reagents required for the staining run brought to room temperature.
14. Select the PRINT button to print the Reagent Layout Map. 15. Load reagents using the Reagent Layout Map. 16. Select the NEXT button. The SLIDE LAYOUT MAP:LOAD SLIDES screen appears. 17. Load slides using the Slide Layout Map. It is recommended that each slide be flooded gently with buffer after it is placed on the instrument. This buffer should contain 0.05% Tween 20. (See Loading Slides, Section 13) At no time should the slides dry out. 18. Select the NEXT button.
Section 5 | Programming Overview The Dako Autostainer is a computer controlled system. The PROGRAMMING GRID is the main screen used to define slide information, primary antibodies, staining reagents and staining protocols. Menu Bar Grid Header Programming Grid Program Name Command Buttons Software Version Date/Time User Name (security level) Institution Name Autostainer serial number 1. Five menus are accessible from the menu bar.
Menu Bar Options Slides Function The Slides option may be used as an alternative to expedite programming by reducing the amount of information required to program a staining run. The Slides option may be used to enter the total number of slides for the current staining run. The Slides function can be accessed by moving the mouse pointer to Slides on the menu bar and pressing the left mouse button. This selection displays the SLIDE COUNT window for entering the number of slides for the current staining run.
Edit Lists Menu The Edit Lists menu can be accessed by moving the mouse pointer to Edit lists on the menu bar and pressing the left mouse button. For instruction on the features of the Edit List menu item, see Reagent Management, section 10. The Edit Lists menu commands include: End. Enz.
Auto Function The Auto function can be accessed by moving the mouse pointer to Auto on the menu bar and pressing the left mouse button. This function is used to save all or any portion of a protocol to be used in other programs. (See the Auto Programming section of Programming Slides, section 10) The Help function is also accessible from this menu.
Section 6 | Accessing the Dako Autostainer Software The SIGN IN screen is the first screen that appears after the Dako Autostainer icon is double clicked. The SIGN IN screen allows you to enter your assigned user name (up to 28 alpha-numeric characters) and password (up to 8 alpha-numeric characters). Your user security status (programmed in the INITIALIZE screen) determines the functions you are able to access. If the sign-in process is executed correctly (i.e.
NOTE: In the example below, the INITIALIZE button is displayed, indicating this user is set at high security level. NOTE: The software version number, current date and time, user name, security level (indicated by asterisks), institution name, and Autostainer serial number are displayed across the bottom of this screen. NOTE: A cleaning run is recommended after 150 slides have been processed on the Autostainer.
Section 7 | Initializing the Dako Autostainer The INITIALIZE screen is used to establish and update default information for the system. This includes the institution information, Autostainer serial number, user names, the doctors requesting IHC tests, a library of tissues that may be used during staining, the number of slides allowed between routine cleaning runs, the default volume and the drop zone for reagent dispense.
The buttons on the INITIALIZE screen and their respective functions are: SLIDE LABELS REAGENT LABELS OPTIONS OK CANCEL PRINT HELP displays the DESIGN SLIDE LABEL screen used to define slide label printing of slide labels. displays the DESIGN REAGENT LABEL screen used to define the printing of reagent labels. displays the OPTIONS screen used to define the format for the Program Grid, Slide Layout Map, and printing format for IHC Reports. saves the entered information and returns to the MAIN MENU screen.
Editing Institution Information 1. Select INITIALIZE on the MAIN MENU screen. The INITIALIZE screen is displayed with the cursor in the Name box and the previously entered information listed. 2. Select the information to be changed and enter the new information. 3. Select the OK button to save the entered information. The MAIN MENU screen is displayed. Select the CANCEL button to escape without saving the changes. The MAIN MENU screen is displayed. Entering Staff Information Adding a New User 1.
5. Type in the same password again and press ENTER. The High security level option becomes highlighted. If a different password is entered the window “TRY AGAIN: Password must be identical” appears. 6. Using the mouse, select the desired security level. A mark appears in the raised diamond. Selecting the High Level option allows full access to the software features. Select OK and the INITIALIZE screen appears.
Deleting User Information 1. Select INITIALIZE on the MAIN MENU screen. The INITIALIZE screen is displayed with the cursor in the Name box and the previously entered information listed. 2. Press ENTER or TAB key until the cursor flashes in the Users box. 3. Select the user name from the list and press ENTER. The PASSWORD AND SECURITY LEVEL screen appears. The DELETE USER button located to the right of the entry boxes becomes available. 4. Select the DELETE USER button.
Deleting Doctor Information 1. Select INITIALIZE on the MAIN MENU screen. The INITIALIZE screen is displayed with the cursor in the Name box and the previously entered information listed. 2. Press ENTER or TAB key until the cursor flashes in the Doctors box. 3. Select a doctor from the current list of doctors. 4. Press ENTER. A dialog box appears asking if you want to delete the selected doctor. 5. Select the YES button. The selected doctor is deleted from the current list and the INITIALIZE screen appears.
Entering Default Reagent Information Assigning Default Reagent Dispense Volumes 1. Select INITIALIZE on the MAIN MENU screen. The INITIALIZE screen is displayed. 2. Select a default reagent volume by clicking on a raised diamond next to the appropriate volume. A smaller black diamond appears next to the selected volume. 3. Select the OK button to save. The MAIN MENU screen is displayed. 4. Select the CANCEL button to escape without saving. The MAIN MENU screen is displayed.
Cleaning Schedule This selection determines the number of slides the instrument will stain prior to prompting the user to start a cleaning cycle. Dako recommends setting the cleaning schedule to 150 slides. The MAIN MENU screen always displays the total number of slides processed since the last cleaning cycle in red. 1. Move and click the mouse pointer on the Slides/Cleaning box and type in the number of slides to stain between maintenance cleanings.
for each field is the data entered in the SLIDE INFORMATION screen and appearance of the Programming Grid. NOTE: If Slide ID, Doctor or Tissue is selected to print on the label, then these items must be selected from the OPTIONS button in the INITIALIZE screen. If these Program Grid items are not selected from the OPTIONS button in the INITIALIZE screen, then these fields will not be present in the SLIDE INFORMATION screen. 2.
3. Select the desired option for the field. The list box disappears and the selected option is displayed in the field. NOTE: Static information can be entered into the field. This information is printed on every label. For example, the institution name can be printed on every label. To enter static information, highlight the text in the desired field and type the text in the text box. Press ENTER. 4.
NOTE: The communication port for the printer is identified using the Port box on the DESIGN SLIDE LABEL screen. The number displayed in the box must match the computer serial port where the label printer is connected. 2. Select the OK button to save. The INITIALIZE screen is displayed. Select the CANCEL button to escape without saving. The INITIALIZE screen is displayed. Printer Selection 1.
2. Select the items to be included on the Program Grid. Any or all of the listed options can be marked. Only checked items are displayed on the SLIDE INFO screen and the Programming Grid. NOTE: Selecting the Comment item creates a column for data entry on the Programming Grid printout. This column is not displayed on the PROGRAMMING GRID. NOTE: Slide ID, Block ID, Doctor, and Tissue items were selected to create this Programming Grid example.
Delayed Start Water Rinse: For a delayed start, idle rinses may be performed with water or buffer. Select the Delayed Start water rinse option to use water. If this option is not checked then idle rinse will be with buffer. (Deionized water is not available with the “01” hardware version of the Dako Autostainer.) NOTE: The Delayed Start Water Rinse options cannot be selected while the No Pre-Rinse option is selected.
This page intentionally blank.
Section 8 | Entering Slide Information Slides can be programmed from the PROGRAMMING GRID either through the Slides function for quick programming of total slide count, or through the SLIDE INFORMATION button for entering specific slide identification information. This section describes both options for programming slides. Slides Function Adding Slides The Slides option expedites programming by reducing the amount of information required to program a staining run. 1.
Deleting a Slide When using the Slides option to program a staining run, a slide can be deleted if there is at least one entirely blank (unprogrammed) row. 1. Select the Slides menu item. 2. Adjust the number of slides and click OK. NOTE: If all rows have a programmed tile the slide must be deleted from the PROGRAMMING GRID (see Deleting a Specific Slide, later in this section.) The Slide Information Button The SLIDE INFO button enables the entry of specific information for every slide to be stained.
The buttons on the SLIDE INFORMATION screen and their functions are: FINISH ENTRY DELETE CANCEL HELP saves the entered information and returns to the Programming Grid. deletes selected programmed cases for the selected Slide Identification or Case #. cancels any entries made and returns to the Programming Grid. displays information to aid in understanding and utilizing the features of the SLIDE INFORMATION screen. Entering Slide Information Through the SLIDE INFORMATION Button 1.
To select a Tissue that was entered at the INITIALIZE screen, press the DOWN arrow key on the keyboard until the tissue appears highlighted in the Tissue box and press ENTER. If the tissue is not listed, it can be typed in the box followed by pressing the ENTER key. The cursor moves to the # Slides box. 10. Type in the number of slides for the case and press TAB or ENTER. The cursor returns to the Block ID box. Enter next block if necessary.
6. Select the FINISH ENTRY button to save the changes. The PROGRAMMING GRID is displayed. Select the CANCEL button to undo any changes made to the slide identification record and the PROGRAMMING GRID is displayed. Deleting a Case # This method deletes all slides associated with the selected slide identification’s case. 1. Select the SLIDE INFO button on the PROGRAMMING GRID. The SLIDE INFORMATION screen is displayed. 2.
9. Select the FINISH ENTRY button to save changes. The PROGRAMMING GRID is displayed. Select the CANCEL button to undo any changes made to the slide identification record and the PROGRAMMING GRID is displayed. Deleting a Specific Slide This method deletes one slide at a time. 1. On the PROGRAMMING GRID select the slide number on the row representing the slide. A dialog box asks if you want to delete the selected slide. Slide # 2.
Section 9 | Designing a Protocol Protocol templates are defined as a sequential series of steps required to stain a microscope slide. Each template is built by selecting the individual protocol steps listed under the Protocol Elements column in the correct order to complete a staining protocol. Protocol templates can be saved for future use. The PROTOCOL TEMPLATE DESIGN screen enables you to create a staining protocol template. An unlimited number of protocol templates can be saved for future use.
Protocol Elements The Protocol Elements are categories of reagents used to build the Protocol Outline in the PROTOCOL TEMPLATE DESIGN screen. Reagents can be added, deleted and edited by choosing Protocol Element categories from the Edit Lists menu item on the PROGRAMMING GRID.
Substrate-Batch This step splits the staining run into two distinct operations. The first includes all steps prior to the application of the substrate and the second batches all steps starting with the substrate application. This feature allows you to use unstable substrates that need to be prepared immediately prior to application. The Autostainer stops after the steps in the first operation (nonSubstrate protocol steps) are completed.
5. Additional steps can be inserted in the Protocol Outline by highlighting a step in this column and then clicking on the desired step from the Protocol Elements. The selected step will be inserted above the highlighted area in the Protocol Outline. NOTE: The Dako Autostainer can accommodate protocol templates containing as many as 35 steps (including rinse steps between reagent incubations). Saving a Protocol Template NOTE: Low level security does not allow access to saving a Protocol Template.
NOTE: When assigning a new template to an existing programmed staining run, a dialog box appears if the new template lacks protocol elements present in the previous template. Select the YES button and the columns of missing protocol elements disappear from the grid while newly programmed columns appear. Select the NO button and the PROGRAMMING GRID appears unchanged. Deleting a Protocol Template NOTE: Low level security does not allow access to deleting Protocol Templates. 1.
Using a New Protocol Template Without Saving It 1. After a Protocol Template has been created or modified, click on the USE TEMPLATE button in the PROTOCOL TEMPLATE DESIGN screen. A dialog box asking “Save on disk now?” is displayed. 2. Select the NO button and the PROGRAMMING GRID is displayed with the new protocol template in the header row.
5. Press ENTER or click on the OK button. The PROTOCOL TEMPLATE DESIGN screen appears with the selected Auto Program protocol template displayed in the Protocol Outline column. 6. Click on the USE TEMPLATE button. The PROGRAMMING GRID appears with the selected protocol template displayed in the grid header. Selecting and Editing the Reagent Dispense Volume NOTE: Low security level does not allow access to changing the reagent volumes for a Protocol Template.
Selecting and Editing the Reagent Dispense Volume for a Specific Step in a Protocol 1. Select a Protocol Template Step in the Protocol Outline column to assign a new reagent dispense volume. The selected step will be highlighted. 2. Select the REAGENT VOLUME button in the upper right corner of the PROTOCOL TEMPLATE DESIGN screen. A dialog box with the dispense volume choices for the selected reagent step appears. 3.
Section 10 | Programming Slides The Autostainer can process a maximum of 48 microscope slides in a single staining run. Each slide can be stained using an independently designed protocol. After the slide information is entered and protocol template is selected, specific reagents are assigned to each slide. This is accomplished from the PROGRAMMING GRID. The following section describes how to program the specific reagents for a staining run.
2. Select the desired reagent from the list using the UP and DOWN arrow keys on the keyboard, the mouse pointer, the reagent list scroll bar or by typing the name of the reagent. A dialog box prompts “Assign to the following unprogrammed slides?”. Select YES to assign the selected reagent to the remaining slides in the grid and the next programmable step is highlighted. Select NO and the next programmable step is highlighted without assigning the reagent to the remaining slides. 3.
Assigning Positive and Negative Control Reagents Positive and negative control reagents can be automatically added for each primary antibody that is added to the Programming Grid. The controls will be added as new slides and will contain the matching detection reagents of the test specimen. 1. Enter Slide Information and select a protocol template. A minimum of one slide must be programmed through SLIDE COUNT or the SLIDE INFORMATION screen. Press OK or the FINISH ENTRY button.
2. In a rinse column, position the mouse pointer over a tile with a (blue) rinse buffer icon and click the right mouse button. This will change the buffer rinse to a blow step. To change the blow step back to a buffer rinse, position the mouse pointer over the blow icon and left click the mouse button. 3. In a rinse column, position the mouse pointer over a tile with a (blue) rinse buffer icon and hold down the left mouse button and click the right mouse button. Release both buttons.
3. Select Auto from the menu bar on the PROGRAMMING GRID using the mouse pointer. 4. Select Setup from the Auto menu. Auto changes to Setup on the menu bar. 5. Select one or more rows of programmed tiles by clicking on the first row. Move the mouse pointer over the desired rows while holding down the left mouse button. Each selected row is high-lighted. Release the mouse button and the Save Auto Program window is displayed. NOTE: An Auto Program can consist of one or more rows of programming. 6.
5. Click on the DELETE PROGRAM button. Select the YES button to delete the Auto Program and return to the SAVE AUTO PROGRAM window. Select the NO button to cancel changes and return to the SAVE AUTO PROGRAM window. Using an Auto Programming Item 1. Select Auto from the menu bar in the PROGRAMMING GRID. 2. Select Program from the Auto menu. The word Auto changes to Program. 3. Select a tile in the row where the Auto program is to be inserted.
4. Move the mouse pointer to the slide(s) where the items are to be pasted and press the left mouse button. The copied reagents are inserted for the selected slides. NOTE: Copied sections can only be pasted under the same heading and cannot be pasted over areas that are currently highlighted. 5. Select Paste on the main menu to deactivate the Copy function. Copy/Paste Entire Rows 1. Select Copy from the menu bar on the PROGRAMMING GRID. The word Copy changes to Select. 2.
Replacing a reagent with skip step (no reagent added to slide): 1. Move the mouse pointer over the reagent step to be changed and press the left mouse button. The list of available reagents is displayed. 2. Select (none). (none) is displayed in the tile. Editing a reagent in a staining step for one run only: 1. To edit a reagent in a staining step or to use a reagent not listed in the reagent list, click on the step and select the (edit slide) item from the reagent list.
Printing Options The Autostainer software allows the user to print out various reports including information on each program, run, and reagents stored in the Autostainer. Slide labels with program related information and reagent labels with information stored in the software can be created using the software. Slide Labels 1. From the PROGRAMMING GRID, select the PRINT button. The WHICH REPORT? screen appears. 2. Select the SLIDE LABELS button. The PRINT SLIDE LABELS window appears. 3.
3. To print all reagent labels, select the PRINT ALL button. To print only selected reagents, select desired reagents by positioning the mouse pointer over a reagent and clicking the left mouse button. To select more than one reagent, repeat this step. To de-select a selected reagent, position the mouse pointer over the highlighted reagent and click with the left mouse button. The reagent will become un-highlighted, or deselected. Once all desired reagents are highlighted, select the OK button.
Immunohistochemical Report (IHC Report) 1. From the PROGRAMMING GRID, select the Print button. The WHICH REPORT? screen appears. 2. Select the IHC button. The IHC Report can also be printed from the RUN LOG screen after the completion of a run. 3. After a run is completed, a PRINT IHC button appears at the bottom of the screen. 4. Select the PRINT IHC button. The IHC Report format is selected during Autostainer initialization.
Viewing Programmed Slides After the completion of reagent programming, the first SLIDE LAYOUT MAP is displayed. The SLIDE LAYOUT MAP screen is a representation of all 48 slides as they would be viewed facing the front of the Autostainer. The information presented for each slide includes the slide number, the primary antibody, the case number and reagent dispense volume. The dispense volume and drop zone(s) for each slide can be changed on this screen.
Each cell on the screen represents a microscope slide. Cells contain the position number, case number and primary antibody short name. If case number and primary antibody name have not been assigned, the cells contain only the position number.
2. Selecting multiple dispense locations: To assign multiple dispense locations on a slide, press the CTRL key while clicking on the desired dispense locations with the mouse pointer. Multiple yellow highlights appear on the slide identifying the selected dispense locations. NOTE: Each of the selected dispense locations will receive the volume of primary antibody programmed. The maximum total dispense volume per slide is 800 µL.
ALL SLIDES Tool At the upper left corner of the SLIDE LAYOUT MAP:PROGRAM SLIDES screen is an ALL SLIDES tool. This tool allows users to change the dispense pattern and volume for all slides with one click. 1. Select a new reagent dispense location on the slide in the ALL SLIDES tool by clicking on the desired dispense location using the mouse pointer. The yellow highlight appears in the selected location on each programmed slide.
This page intentionally blank.
Section 11 | Reagent Management Reagent lists are defined using the commands on the Edit lists menu on the PROGRAMMING GRID. A new reagent can be created and existing reagents can be modified or deleted from the reagent list using this function. Changes can be made to all reagent-related information (reagent name, abbreviation, compatibility code, lot #, expiration date, and incubation time). The following procedures are applicable to all lists except Primary Antibody (see Notes below).
3. Type the full name of the reagent you want to add to the current list and press TAB or ENTER. The software allows the entry of 33 alpha-numeric characters for detection reagents. No symbols can be used in the reagent long name. Once the box is filled an audible signal alerts that no additional characters can be entered. The cursor moves to the Short Name box. 4. Enter an abbreviated name for the reagent in the short name box and press TAB or ENTER.
Adding Primary Antibodies A primary antibody can be added to the reagent list with a specific pretreatment reagent. This will facilitate automatic selection of the pretreatment whenever a primary antibody is selected at the PROGRAMMING GRID. 1. From the PROGRAMMING GRID select Edit lists from the menu bar. The reagent lists categories are displayed. 2. Select the Primary Antibody category. The Primary Antibody EDIT REAGENT LIST screen is displayed with the cursor in the text box in the Antibody section.
5. If the primary antibody is a step that is preformed off the Autostainer (i.e. overnight incubation or requires heating) then check the Manual Step check box by pressing the SPACE BAR. When this reagent is selected at the PROGRAMMING GRID, “(Manual Step)” is displayed in the reagent tile, signifying that this step is preformed before the slides are loaded on the Autostainer. Press TAB or ENTER. The cursor moves to the Compatibility box. 6. Enter the correct compatibility code and press TAB or ENTER.
13. Enter the reagent incubation time and press TAB or ENTER. NOTE: If Manual Step was selected, the incubation time will be “0”. Do not change this time. The cursor moves to the first text box to allow addition of the next primary antibody Follow steps 3-13 until all antibodies have been added. NOTE: If a new pretreatment was used for this antibody, message appears asking if this pretreatment should be added to the Pretreatment list. Select Yes to add reagent to the pretreatment list. 14.
3. Display the specific reagent by selecting the scroll bar in the Reagent Name box or press the UP or DOWN arrow key on the keyboard until the target reagent appears highlighted. Press ENTER and the reagent information is displayed. 4. Select the DELETE button. A dialog box asks if you want to delete the selected reagent. 5. Select the YES button. The EDIT REAGENT LIST screen appears and the reagent is deleted. Selecting the NO button displays the EDIT REAGENT LIST screen and the reagent is not deleted. 6.
Printing Reagent Labels Printing reagent labels from a programmed run. 1. Program or retrieve a previously saved run on the PROGRAMMING GRID, select the PRINT button. The WHICH REPORT? screen appears. 2. Select REAGENT LABELS button. The PRINT REAGENT LABELS screen appears displaying all reagents from the programmed run on the PROGRAMMING GRID. 3. To print all reagent labels, select the PRINT ALL button.
Compatibility Check A compatibility check has been built into the Dako Autostainer software to minimize programming errors resulting from assigning incompatible reagents (such as alkaline phosphatase-labeled streptavidin and DAB substrate) to a slide protocol. This feature has two check levels. The first check is the species reactivity compatibility of the primary antibody and the secondary reagent used. The second addresses the compatibility rules dictated by the enzyme used in the detection system.
Every reagent in the reagent categories lists (Protocol Element) can be programmed with an appropriate compatibility code or combination of codes. For example a linking reagent that reacts with mouse and rabbit can be marked with compatibility codes AB. The software verifies that these compatibility rules are not violated during the programming of a staining run.
This page intentionally blank.
Section 12 | Loading Reagents After programming is completed the Autostainer calculates the time required for the staining run, the amount of wash buffer/deionized water required and the most efficient placement for the reagent vials. 1. Select the NEXT button in the SLIDE LAYOUT MAP screen. A dialog box signaling that the calculation is complete lists the run time, number of probe washes and the volume of buffer and deionized water required.
If more than 32 reagent vials are required by the run, a SECOND RACK button becomes available at the bottom of the screen. The buttons and their functions are: REAGENT LIST NEXT BACK TO GRID SLIDE MAP FIRST/ SECOND RACK PRINT displays a detailed listing of the reagents used in the current staining run. displays the SLIDE LAYOUT MAP:LOAD SLIDES screen. returns to the Programming Grid. displays the SLIDE LAYOUT MAP:Review Slides screen.
Missing Reagent Error The Dako Autostainer prompts the user when an insufficient volume of reagent is measured for a run. The Autostainer can be paused to allow the user to add more reagent to the specified reagent vial. If sufficient volume of a reagent is not available, the Autostainer will provide a list on the Run Log of skipped or partially applied slides.
Partial Reagent Application Reagent vials are loaded on the Autostainer and a run has been started. 1. The reagent probe senses insufficient or a missing reagent. The MORE REAGENT window is displayed on the screen. The screen states the reagent vial position, amount needed and reagent that is missing. 2. A 60 second count begins to count down and an alarm sounds. 3. If the user does not notice the MORE REAGENT alarm, after 60 seconds the RUN LOG screen appears and the pause in the run is recorded. 4.
Section 13 | Loading Slides After programming is completed and the reagents have been loaded, you are ready to load the slides on the slide racks. Print the Slide Layout Map by selecting the PRINT button in the SLIDE LAYOUT MAP:LOAD SLIDES screen. This map provides a guide for loading slides and can be saved as part of your staining record. Loading Microscope Slides NOTE: The slides should soak in a buffer solution containing 0.05% Tween 20 for a minimum of 5 minutes prior to loading on the Autostainer. 1.
This page intentionally blank.
Section 14 | Starting a Run After the specimen-containing slides and reagent vials have been loaded, it is advisable to go through the following checklist: Slides • • should be flooded with buffer containing 0.05% Tween 20 using a squirt bottle to prevent slides from drying while loading additional slides. should be seated securely in slide rack. Racks • should be properly seated in the Autostainer. Reagents • • • • should be at room temperature.
Starting a Run without Delay Starting a staining run immediately. 1. Select the NEXT button on the SLIDE LAYOUT MAP:LOAD SLIDES screen during slide loading. The SET START TIME window is displayed. The Start Time is listed as +0:00 and the current time. The Finish Time is listed as the +HH:MM required to complete the run. The calculated finish time is also displayed. 2. Select the PRIME PUMP (Water) button. A dialog box appears asking if the pathway is clear for the arm to move. Select the YES button.
5. Select the START RUN button. The RUN LOG screen is displayed. The Run Log is visible throughout the run and displays the steps performed by the Dako Autostainer. The Elapsed, Remaining, Total, Start, Current, and Finish times are listed on the RUN LOG screen. The current step, total and current time are always displayed in blue at the bottom of the Run Log. NOTE: If the current time is not accurate do not change it during the run. Adjust the time at the completion of the run.
Delaying the Start of a Run Starting a staining run at a later time. 1. Select the gray scroll bar and hold the left mouse button down. Move the scroll bar to the right until the desired Start Time and Finish Time is displayed. Release the mouse button. The staining run will start and finish at the times indicated. Gray scroll bar used to adjust start time in large increments. Scroll arrow used to adjust start time in small increments.
4. Select the START RUN button. The RUN LOG screen is displayed. The Run Log is visible throughout the run and displays the steps performed by the Autostainer . The elapsed, remaining and total times are listed on the RUN LOG screen. The Start, Current, and Finish times are also listed. The current step, total and current time are always displayed in blue at the bottom of the Run Log. NOTE: If the current time is not accurate do not change it during the run. Adjust the time at the completion of the run.
2. Select the START RUN button. The RUN LOG screen is displayed. The elapsed, remaining and total times are listed on the RUN LOG screen. The Start, Current, Finish, and Batch times are also listed. The current step, total and current time are always displayed in blue at the bottom of the Run Log. NOTE: If the current time is not accurate do not change it during the run. Adjust the time at the completion of the run. To reset time refer to the Windows 95/98 documentation.
Section 15 | Completing a Staining Run During a staining run the RUN LOG screen is displayed and updated with each step performed. The current step is highlighted in blue and displayed at the bottom of the screen. Previously performed steps can be reviewed using the scroll bars. NOTE: To view previous steps, use the scroll bar to get to the desired view field, then release the mouse button. Do not hold the button depressed for an extended period.
NOTE: If the Print IHC button is not pressed, then a Run Log Report will not be printed. To print out an IHC report at a later time, see Programming Slides, section 10. 2. Select the OK button. The robotic arm moves to the home position (left side of the instrument) and the waste pump activates to drain the waste reservoir. A dialog box asks if the Run Log should be saved. For instructions on how to save and print a run log see Saving and Printing a Run Log below. Select YES to save the Run Log.
Printing a Temporary Run Log If the Autostainer comes to an unexpected stop (i.e. power failure), a temporary copy of the Run Log is maintained on the computer. This record can be retrieved and the current step of each slide can be identified so that a run can be restarted. 1. Restart the computer. NOTE: If the Autostainer was not shut down properly, then a Scan Disk may occur when you start up the computer. Follow the instructions on the screen if a Scan Disk occurs. 2.
NOTE: If the Autostainer is stopped when there is reagent in the probe, it will ask you if you would like to clean the probe. Select yes if it is safe to allow the arm to move. If there is as object in the pathway of the arm or if the arm has been moved out of calibration during the run, select No. NOTE: If the ESC key is pressed during Autostainer operation, this will emergency stop the run. Select the YES button to stop the run. Select the NO button if you do not want to abort the run.
Section 16 | Programming During Staining Run This feature allows the user to program subsequent runs while the Autostainer is in operation. All programming screens are accessible, including the PROGRAMMING GRID, SLIDE LAYOUT MAP screen, RUN TIME CALCULATION screen and REAGENT LAYOUT MAP screen. A print out of all screens can be generated after the current run is completed. The Run Log screen displays the features described below.
Reviewing the Current Program during a Staining Run This feature allows the user to review the PROGRAMMING GRID for the current run while the Autostainer is in operation. No changes can be made using this function. While the current run is in process and the RUN LOG screen is displayed: 1. Select the REVIEW PROGRAM button on the RUN LOG screen. The PROGRAMMING GRID appears with the programming grid of the current run displayed. 2. To return to the RUN LOG screen select the NEXT or EXIT button.
Section 17 | System Maintenance It is recommended that the Dako Autostainer be cleaned after 150 slides have been processed on the instrument. The cleaning procedure can be divided into a manual phase and an automated phase. Manual cleaning consists of wiping down any areas exposed to reagents and buffer using deionized (DI) water and a detergent. Organic solutions are not recommended for cleaning.
NOTE: If running protocols that require more frequent cleaning (i.e. using hematoxylin counterstain or high salt buffers), Dako recommends performing a cleaning run a the end of each day the instrument is used. Cleaning Log Each time the Autostainer is cleaned, this is logged in a Cleaning Log so that a record of each cleaning is maintained. Each time a cleaning is reset without performing an actual cleaning run, this is recorded in the Cleaning Log with the notation “(Reset)”. 1.
Slide Rack Cleaning Dako recommends additional cleaning for slide racks. 1. Soak the slide racks overnight in a container of deionized water. 2. After soaking the racks, rinse with deionized water and brush off any salt residue before beginning the next run. Reagent Vial Cleaning The cleaning and re-use of reagent vials is not recommended by Dako. Additional vials can be purchased from Dako (Code No. S3425). Computer Maintenance Follow instructions as defined by the computer operator’s manual.
This page intentionally blank.
Section 18 | Reagent Tracking Reagent Tracking allows you to monitor the total volume of each reagent that has been used on the Autostainer. A Dako Autostainer Tracking Report can be generated to display various information about all the reagents used on the instrument. After each Autostainer run, the usage of each reagent is tracked on the Reagent Tracking screen.
Remove Reagent from the Reagent Tracking Log Reagents that are outdated or not critical to tracking can be removed from the Reagent Tracking log. Remove all Reagents from the Reagent Tracking Log 1. Select the RESET ALL button. A RESETTING REAGENT window appears and asks if you are sure you want to reset all reagents. 2. Select YES to remove all reagents from the Reagent Tracking Log. Select NO to cancel this action and return to the REAGENT TRACKING screen.
Section 19 | Wash Buffers Wash buffers are required for all immunohistochemical staining procedures. They function to eliminate background by washing away the nonspecifically bound proteins from the specimen. Commonly used buffers for immunohistochemical staining are Tris-buffered saline (TBS) or phosphate buffered saline (PBS). These buffers are recommended by Dako for use on the Dako Autostainer. The following pages describe detailed instructions for preparation of TBS and PBS buffer solutions.
Reagent Preparation One Liter of Tris-Buffered Saline Solution (S3001) 1. Add 1L (1000 mL) of deionized water to a 1L carboy, flask or beaker. 2. Empty entire contents of one packet. Use a magnetic stir plate to mix and dissolve the TBS granules. 3. Add 0.5 mL of Dako Tween 20 (Code No. S1966) to the Tris-buffered saline solution and mix using a magnetic stir plate. 4. The final solution is 50mM Tris-HCl, 150mM NaCl buffered saline, pH 7.6 with 0.05% Tween 20.
Shelf Life and Storage TBS solutions should be stored at 2-8°C to inhibit bacterial growth. TBS solutions that become turbid should be discarded. If Tris-buffered saline is stored at 4°C, the expected shelf life is 7 days. If Tris-buffered saline is stored at room temperature, the expected shelf life is 4 days. NOTE: Dako Autostainer Wash Buffer does not contain any preservatives.
6. The final solution is 0.5M Tris-HCl, 1.5M NaCl, buffered saline, pH 7.6 with 0.5% Tween 20. One Liter of Tris-Buffered Saline, containing 0.05% Tween 20 from 10x stock 1. Add 900 mL of deionized water to a 1L (1000 mL) carboy, flask or beaker. 2. Add 100 mL of 10X Tris-buffered saline solution and mix using a magnetic stir plate. 3. The final solution is 50mM Tris-HCl, 150mM NaCl buffered saline, pH 7.6 with 0.05% Tween 20. One Liter Tris-Buffered Saline containing 0.05% Tween 20 1.
Dako Phosphate Buffered Saline Dako Phosphate Buffered Saline Solution can be used for Autostainer reagent preparation and general laboratory use. Intended Use Dako recommends phosphate buffered saline (PBS) in the preparation of many immunohistochemical and in situ hybridization procedures. Materials Required 1. Clean carboy, flask or beaker graduated to 1 Liter 2. Deionized water 3. Magnetic stir plate with stir bar 4. Prepackaged Dako reagents Code No.
Phosphate Buffered Saline The following is a procedure for the formulation of 20mM Phosphate, 150mM NaCl, buffered saline, pH 7.0 with 0.05% Tween 20 from common laboratory chemicals. Intended Use A phosphate buffered saline solution, pH 7.0 can be used on the Dako Autostainer. Materials Required 1. Clean carboy, flask or beaker graduated to 1L 2. Deionized water 3. Magnetic stir plate with stir bar Common laboratory chemicals Chemical Manufacturer and Product No.
Shelf Life and Storage PBS solutions should be stored at 2-8°C to inhibit bacterial growth. PBS solutions that become turbid should be discarded. If phosphate buffered saline is stored at 4°C, the expected shelf life is 7 days. If phosphate buffered saline is stored at room temperature, the expected shelf life is 4 days. NOTE: The above formulation for phosphate buffered saline does not contain any preservatives.
This page intentionally blank.
Section 20 | Troubleshooting Solving Autostainer Problems Symptom Possible Cause Solutions Autostainer will not turn on. (top green LED not on) A Cannot Continue box displays “Instrument Not On” when a run is started. The Autostainer is unplugged at the power source. Check the power cord connections and verify the surge protector is plugged in and turned on. The Autostainer does not respond when you initiate a prime pump or a run.
Symptom Possible Cause Solutions Buffer or DI Water does not flow out during a run or a prime. (Cont.) Inlet filters are clogged and are restricting fluid flow. Check the inlet filter at the end of the tubing in the carboy clean if necessary. Dako recommends using a tube and cap assembly (S3472) to prevent foreign objects from entering the carboys and possibly clogging the filter. Tubing not completely inserted into carboy.
Symptom Possible Cause Solutions Reagent probe is not dispensing reagent. (Cont.) Clogged reagent probe. Contact your local Dako representative. (See Dako Companies and Distributors, Section 23) Bubbles on the surface of the reagent in the reagent vial. Remove bubbles. The yellow 3- way “stopcock” valve on the syringe has been moved. Verify that the “OFF” position of the valve is in the down direction. The valve should be in a “T” configuration. Arm is out of alignment.
Symptom Possible Cause Solutions No Staining Insufficient reagent in reagent vial. The Autostainer is equipped with a liquid level sensor. If the sensor detects insufficient reagent to complete the current step, a message appears on the screen prompting you to add missing reagent to the correct reagent vial. You have one minute to add reagent after which some slides may be skipped. Check Run Log for “Insufficient Reagent **Skipped Slides**” message.
Solving Software Problems Symptom Possible Cause Solutions User is unable to access many of the functions in the Initialize section of the program. User has been set up as a “Non-Authorized” user. Contact your local Dako representative. (See Dako Companies and Distributors, Section 23) Autostainer software will not accept your log-in name and password. Name and password not entered properly. Ensure that you are using the proper spelling. Illegal or Fatal exception errors. Windows error.
Solving Computer Problems Symptom Possible Cause Solutions The computer will not turn on. The computer is unplugged at the power source. Check the power cord connections and verify the surge protector is plugged in and turned on. Wall outlet is not working. Ensure the wall outlet is operational. Check also for a blown fuse, or try using a different wall outlet for the surge protector. Surge protector is not working. Check to see if the circuit breaker on the surge protector is popped out.
Solving Monitor Problems Symptom Possible Cause Solutions Monitor will not turn on or goes blank intermittently. (power LED is not on) The monitor is unplugged at the power source. Check the power cord connections and verify the surge protector is plugged in and turned on. The monitor screen is not displaying normally. The cable between the monitor and the computer is not connected properly. Disconnect and reconnect the cable between the monitor and the computer. Monitor screen went blank.
Symptom Possible Cause Solutions Image on the desktop is grainy and the color is incorrect. (Cont.) Color depth has been adjusted too low. At the Windows Desktop, use the mouse to right click anywhere other than an icon, a drop down menu will appear. Select Properties. The WINDOWS DISPLAY PROPERTIES window should appear. Select the Settings tab. In the box marked colors verify that “High Color (16 bit)” is displayed. If it is not, select it and save changes.
Solving Printer Problems Symptom Possible Cause Solutions The printer does not respond to print command. The printer may be in Manual Feed mode. Briefly press and release the Front Panel Button on the printer. The parallel cable between the printer and the computer is not connected properly. Disconnect and reconnect the parallel cable between the printer and the computer. The parallel cable is defective.
Symptom Possible Cause Solutions All the printer LEDs are on steadily (not blinking). The printer has a hardware error. Disconnect the printer from the surge protector and wait 1 minute before reconnecting. If all three LEDs come back on, contact your local Dako representative. The printer is not feeding properly or jams frequently. Paper was added to the Paper Input Bin incorrectly. Remove the paper from the Paper Input. Tap the sides of the stack together and reinsert into the bin.
Solving Seymour Label Printer Problems Symptom Possible Cause Solutions The text of the label does not begin along the left edge of the label and the text is getting cut off. The Seymour printer may need to be reset. To reset the Seymour Label Printer, turn off the printer. Hold down the green label feed button on the top of the printer. Turn on the printer while holding down the green button and continue holding the button for 3 seconds. The labels advance forward and printer will print.
This page intentionally blank.
Section 21 | Warnings and Symbols Keep the cover closed during operation. The robotic arm will move unexpectedly during the operation - stay clear. Do not pour liquid down the Dako Autostainer sink. The Dako Autostainer is not equipped to drain large liquid volumes poured at high speed. Do not use bleach in the Dako Autostainer. Bleach may react with other chemicals and create toxic fumes. Do not impair the movement of the Dako Autostainer robotic arm in any way.
Contact your local Dako representative prior to using non-Dako reagents or solutions on your Autostainer. Some solvents, acids, and other solutions may cause damage to internal components of the Autostainer and affect your instrument's performance and warrantee.
Section 22 | General Limitations Immunohistochemistry is a multi-step diagnostic process that requires specialized training in the selection of the appropriate reagents, tissue selection, fixation, and processing, preparation of the IHC slide, and interpretation of the staining results. Tissue staining is dependent on the proper handling and processing of tissues prior to staining.
