Dako Autostainer User Manual

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Dako Autostainer | English Handbook

ii Autostainer Handbook

Dako Autostainer Handbook

Document Number 0000620

Revision A

July 2006

Autostainer Handbook

iii

iv Autostainer Handbook

Copyright © 2006 Dako, Inc. All rights reserved.

This document may not be copied in whole or in part or reproduced in any other media without the express written permission of Dako, Inc. Please note that under copyright law, copying includes translation into another language.

User Resources

For the latest information on Dako products and services, please visit the Dako Web site at:

http://www.dako.com

Installation Procedure

Dako employees will perform the initial installation and setup of all new System instruments.

Relocation Procedure

Contact Dako’s Technical Service Group before relocating your system.

Scope

This handbook contains basic information on the use and operation of the Autostainer and assumes you have received basic training on the instrument. Please contact our Technical Service Group or refer to the Autostainer User Guide for information not provided in this manual. This manual does not provide instructions for the installation or upgrade of hardware.

Disclaimers

This manual is not a substitute for the detailed operator training provided by Dako, Inc., or for other advanced instruction. Dako Technical Service Group should be contacted immediately for assistance in the event of any instrument malfunction. Installation of hardware or software on your Autostainer should be performed only by a certified Dako Field Service Representative.

Contact Information

U.S. Customers

Dako, Inc 6392 Via Real Carpinteria, CA 93013 USA Tel 805-566-6655 Fax 805-566-6688 Technical Support 800-424-0021 Customer Service 800-235-5763

Outside U.S.

Dako Denmark A/S Produktionsvej 42 DK-2600 Glostrup Denmark Tel +45 4485 9500 Fax +45 4485 9595

Autostainer Handbook

vi Autostainer Handbook

Warnings, Precautions and Limitations............................................................................. 1

System Overview .............................................................................................................. 5

Software Overview ............................................................................................................7

Sign In Screen............................................................................................................... 7

Main Menu ....................................................................................................................7

Initialize ......................................................................................................................... 8

Label Printing Initialization ........................................................................................8

Communication Port.................................................................................................. 9

Printer Selection...................................................................................................... 10

The Options Screen ................................................................................................ 10

Format Program Grid ..............................................................................................10

IHC Report ..............................................................................................................10

Custom Options ......................................................................................................10

Reagent Volume and Drop Zone Default ................................................................11

Clean........................................................................................................................... 11

Sign Off and Help........................................................................................................ 11

Reagent Tracking........................................................................................................ 11

Prime Pump ................................................................................................................12

Programming Grid Overview....................................................................................... 12

Entering Slide Information............................................................................................... 13

Adding Slides .............................................................................................................. 13

The Slide Info Button................................................................................................... 13

Deleting Slides ............................................................................................................ 14

Deleting Slides With the Slides Function.................................................................14

Deleting Slide ID Information Using the Slide Info Button....................................... 14

Rearranging slide positions on the Programming Grid ...............................................14

Designing a Protocol....................................................................................................... 15

Standard Protocol Elements .......................................................................................15

Additional Protocol Elements ...................................................................................... 16

Rinse Buffer ............................................................................................................16

Rinse Water ............................................................................................................16

Standard Rinse Replacement ................................................................................. 16

Substrate-Batch ......................................................................................................16

Switch...................................................................................................................... 16

Creating and Editing a Protocol Template ..................................................................16

Deleting a Protocol Template...................................................................................... 17

Using a New Protocol Template Without Saving ........................................................17

Using a Saved Protocol Template............................................................................... 17

Using a Saved Protocol Template for a Specific Auto Program.................................. 18

Selecting Reagent Dispense Volume for All Steps in a Protocol ................................ 18

Selecting Reagent Dispense Volume for a Specific Step in a Protocol ......................18

Programming a Staining Run .......................................................................................... 19

Defining Detection Reagents ......................................................................................19

Defining Primary Antibodies........................................................................................ 20

Assigning Positive and Negative Control Reagents.................................................... 20

Adding Positive and Negative Control Reagents ....................................................21

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Negative Controls.................................................................................................... 21

Defining Specific Rinses .............................................................................................22

Auto Programming ...................................................................................................... 22

Deleting an Auto Programming Item .......................................................................23

Using an Auto Programming Item ...........................................................................23

Copy/Paste.............................................................................................................. 23

Editing a Protocol Step................................................................................................ 24

Printing Options........................................................................................................... 24

Slide Labels............................................................................................................. 24

Reagent Labels .......................................................................................................25

Reports.................................................................................................................... 25

Viewing Programmed Slides .......................................................................................25

Assigning Reagent Dispense Locations.................................................................. 26

Printing a Slide Layout Map ....................................................................................26

Selecting a new reagent-dispense location for all slides:........................................ 26

Reagent Management..................................................................................................... 27

Adding Detection Reagents (Secondary, Tertiary, Substrate, etc.) ............................27

Adding Primary Antibodies.......................................................................................... 28

Editing and Deleting Reagent Lists .............................................................................28

Updating a Reagent Lot Number and Expiration Date................................................ 28

Compatibility Check ....................................................................................................29

Loading Reagents........................................................................................................... 31

Reagent Layout Map Screen ......................................................................................31

Missing Reagent Notice .............................................................................................. 32

Loading Slides.................................................................................................................33

Load Slides .................................................................................................................33

Starting a Staining Run ...................................................................................................35

Preparing the System for a Staining Run.................................................................... 35

Quick Start ..................................................................................................................35

Run Log Screen ...................................................................................................... 36

Shutting Down the Autostainer.................................................................................... 36

Maintenance and Troubleshooting.................................................................................. 37

Troubleshooting ..........................................................................................................37

System Specifications ..................................................................................................... 43

Hardware Specifications .............................................................................................43

Software Specifications............................................................................................... 43

Standard Configuration ...................................................................................................45

viii Autostainer Handbook

Section 1

Warnings, Precautions and Limitations

Keep the cover closed during operation. The robotic arm will move unexpectedly during the operation - stay clear. Do not impair the movement of the Autostainer robotic arm in any way.

Do not pour liquid down the Autostainer sink. The system is not equipped to drain large liquid volumes poured at high speed. Hazardous reagent wastes must be disposed of according to local, state, and federal regulations. Wear appropriate personal protective equipment to prevent exposure.

Do not use bleach in the Autostainer. Bleach may react with other chemicals and create toxic fumes.

Do not attempt to service the Autostainer unless instructed to do so by an authorized Dako representative. Doing so will void the warranty or service contract. Do not relocate the Autostainer System within your facility before contacting your local Dako representative for vital information that may affect your warranty.

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 warranty.

Remove caps from reagent vials before starting a run on the system. Place the reagent racks firmly in their seated position before starting a run.

Place the slide racks in the down position before starting a run on the system.

Do not use the scroll bars to select reagents from the reagent lists when setting up subsequent runs while the Autostainer is processing. Using the scroll bars during operation will cause the Autostainer to pause temporarily.

Do not use symbols when programming reagents or protocols. Doing so may cause errors during the run.

Do not run more than one software application during operation (this includes the CD player and screen savers). Do not install third-party software or hardware products. Installing third-party products may lock up the Autostainer and may void the warranty.

Autostainer Handbook

Do not print slide labels while the Autostainer is running.

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. Improper fixation, freezing, thawing, washing, drying, heating, sectioning or contamination with other tissues or fluids may produce artifacts, antibody trapping, or false-negative results. Inconsistent results may be due to variations in fixation and embedding methods, or to inherent irregularities within the tissue. Excessive or incomplete counterstaining may compromise proper results.

Use of old or unbuffered fixatives, or exposure of tissues to excessive heat (greater than 60°C) during processing may result in decreased staining sensitivity.

Normal/non-immune sera from the same animal source as the secondary antisera used in blocking steps may cause false-negative or false-positive results due to autoantibodies or natural antibodies. False-positive results may be seen due to nonimmunologic binding of reagents to tissue sections. In some case the application of an alternate blocking reagent prior to incubation with the primary antibody may be useful for reducing background. A recommended blocking reagent is Dako Protein Block SerumFree (Code No. X0909).

Unexpected negative reactions in poorly differentiated neoplasms may be due to loss or marked decrease of antigen expression or nonsense mutation in the gene(s) coding for the antigen. Unexpected positive staining in tumors may be from expression of an antigen not usually expressed in morphologically similar normal cells, or from persistence or acquisition of an antigen in a neoplasm that develops morphologic and immunohistochemical features associated with another cell lineage (divergent differentiation). Histopathologic classification of tumors is not an exact science and some literature reports of unexpected staining may be controversial.

The clinical interpretation of any positive staining or its absence should be complemented by morphological and histological studies with proper controls. Evaluations should be made within the context of the patient’s clinical history and other diagnostic tests. It is the responsibility of a qualified pathologist who is familiar with the antibodies, reagents and methods used to interpret the stained preparation. Staining must be performed in a certified licensed laboratory under the supervision of a pathologist who is responsible for reviewing the stained slides and assuring the adequacy of positive and negative controls.

Reagents may demonstrate unexpected reactions in previously untested tissues. The possibility of unexpected reactions in tested tissue groups cannot be completely eliminated due to biological variability of antigen expression in neoplasms, or other pathological tissues. Contact your local Dako representative with documented unexpected reactions. Tissues from people infected with hepatitis B virus and containing hepatitis B surface antigen (HBsAg) may exhibit nonspecific staining with horseradish peroxidase.

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Correct Disposal of this Product (according to Directive 2002/96/EC on Waste Electrical and Electronic Equipment [WEEE] applicable in the European Union and other European countries with separate collection systems).

Contact a Dako representative for disposal of the equipment at the end of its working life. This product should not be mixed with other commercial waste for disposal.

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4 Autostainer Handbook

Section 2

System Overview

The Autostainer System is an automated slide processing system compatible with currently available reagents for staining 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.

Flexible programming allows for an unlimited number of protocols containing up to 35 steps (including rinse and blow steps between reagent incubations) and 64 different reagents. A staining run can process from 1 to 48 microscope slides. Individual slides can be programmed to receive different reagents, of specified volume, during any step in a staining protocol, and waste is segregated into hazardous and non-hazardous collection containers, reducing disposal costs.

The Autostainer is designed to track a variety of data. It can generate patient, reagent, and real-time operation data reports, as well as track reagent usage and log instrument maintenance.

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Section 3

Software Overview

Sign In Screen

The Sign In screen appears when you double-click the Autostainer icon on the desktop. After your name and password have been validated, the Main Menu appears.

Main Menu

The buttons on the Main Menu allow you to access the other Autostainer screens. Note: Change Password replaces the Initialize button for users with middle and low

security access.

Autostainer Handbook

Initialize

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 default volume and the drop zone for reagent dispensation, and the number of slides allowed between routine cleaning runs. Slide and reagent label formatting, user-defined printing formats for the Programming Grid, and IHC Reports can also be configured from this screen.

Label Printing Initialization

The Slide Labels and Reagent Labels buttons are displayed on the Initialize screen and are used to access the Design Label screens for configuring and adjusting slide and reagent label printing.

Slide Labels

The Slide Labels button is displayed on the Initialize screen and it is used to access the Design Slide Label screen for configuring and adjusting slide label printing. Slide labels are printed from the Print button on the Programming Grid. Do not print labels while the Autostainer is operating.

1. Select the Slide Labels button.

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

Programming Grid items are not selected by using the Options button in the Initialize screen, then these fields will not be present in the Slide Information

screen.

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2. Select the desired option for each field.

Note: Static information, such as the institution name, can be entered into the field. This information is 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.

3. Select the number of characters for each line (10, 12, 14, or 16) by selecting a

number from the Length list (Selecting 10 characters will display bold text).

Reagent Labels

Access the Reagent Labels button through the Initialize screen.

1. Select the Reagent Labels button. Reagent Name 1/2 and Reagent Name 2/2 are

used together on adjacent lines to split a long reagent name into 2 separate lines. Reagent Name 1/2 will contain the first half of the reagent name, and Reagent Name 2/2 will contain the second half of the reagent name. The length of each line is set up using the Length tool.

2. Select the desired option for each field.

3. Select the number of characters for each line (10, 12, 14, or 16) by selecting a

number from the Length list (Selecting 10 characters will display bold text).

Label Adjustment

1. To check the position of the text on the slide or reagent label and to generate a

test label, select the Adjust Label button from the Design Slide Label or Design Reagent Label screen.

2. Select the Test Print button. A label is generated by the label printer.

Note: The horizontal adjustment is automatically positioned by the printer. If the text does not begin along the left edge of the label, the printer may need to be reset. Turn off the printer. Hold down the green label feed button on top of the printer and turn the printer on again. When the LED begins to flash red, release the label reed button and then quickly press the button twice. The labels advance forward and printer will print. Release the feed button. Press and release the feed button one more time to advance one label. Perform a test print.

Communication Port

The communication port for the printer is assigned using the Port box on the Design Slide Label screen. The number displayed in the box must correspond to the

communication serial port where the label printer is connected (e.g., 1 for A and 2 for B).

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Printer Selection

To select the proper Seymour printer type on the Design Slide Label screen, scroll though the printer list using the keyboard arrows or the scroll bar.

Note: If the incorrect printer is selected, error messages display on the screen when slide or reagent labels are printed. To find the model number of your Seymour Glass Label printer, check the cover of the printer for a number (e.g.,TPL2642). The last four digits represent the model number (e.g., 2642).

The Options Screen

The Options button is displayed on the Initialize screen. The Options screen enables you to select format options for the Programming Grid and IHC Report, in addition to options for rinsing, and slide layout maps.

Format Program Grid

This function enables you to select format options for the Programming Grid. Selected 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

screen.

IHC Report

Printing options for the IHC Report include:

• One slide ID/page – prints each Slide ID on a separate page

• One case/page – prints one case per page

• No page break – prints continuously all slides for a run

Custom Options

This function enables you to select rinse and blow options and modify the Slide Layout Map. To activate an option click the corresponding check box.

Delayed Start Water Rinse:

For a delayed start, idle rinses may be performed with water or buffer. If this option is not selected, buffer will be used for the idle rinse. Note: This option cannot be selected when the No Pre-Rinse option is selected.

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Add Rinse to Protocol Steps:

When creating a new protocol template, a buffer rinse can be added automatically, following each step. Select the Add Rinse to Protocol Steps option to automatically add buffer rinses. If this option is not checked, the rinse step must be manually selected.

No Pre-Rinse:

When creating a new protocol template, the default pre-rinse step may be removed. If this option is not checked, the protocol template will include a buffer rinse step.

Flex-time Blow:

When a blow instead of a rinse is required, this option enables Autostainer to perform the blow without interrupting other functions.

Show Only Programmed Slides: This option displays only the programmed slides on the Slide Layout Map.

Reagent Volume and Drop Zone Default

From the Initialize screen you can set the default reagent volume by clicking a raised diamond next to the appropriate volume. Select the default drop zone by clicking the desired area of the slide image. Select multiple drop zones by holding the CTRL key while clicking drop-zone areas.

Clean

The Clean screen allows you to initiate a cleaning run. It gives a status report on the number of slides processed since the last cleaning cycle and the total number of runs. It also generates a cleaning log.

A cleaning run is recommended after 150 slides have been processed or after one week, whichever comes first. Once the instrument has processed 150 slides, a message prompts the user to clean the instrument. The software allows up to 200 slides to be run between cleaning cycles to allow for flexibility in workflow. If more than 200 slides have been run, the Program button becomes unavailable and the system does not allow additional processing until the instrument is cleaned.

Sign Off and Help

Sign Off exits the program and Help displays information regarding Main Menu

features.

Reagent Tracking

The Reagent Tracking screen allows you to monitor the total volume of each reagent used by the instrument, and generate reports that include all or specific reagents.

Autostainer Handbook

Prime Pump

The Prime Pump screen is used to flush the intake tubing with buffer or DI water without initiating an actual run. Frequently running fluid through the instrument keeps the Autostainer running properly. Dako recommends priming the pumps weekly.

Programming Grid Overview

The Programming Grid is the main screen used to define slide information, primary antibodies, staining reagents, and staining protocols.

Programming Grid

rows represent individual slides including complete identification and all reagents applied. The grid can accommodate 48 slides per staining

Software Version

Menu Bar

Slides – enter the number of slides for a run File – general information such as opening, saving, and printing slide runs Edit lists – establish lists of reagents for programming a staining protocol Copy and Auto – items used during programming

Date/Time

User Name ** high security * medium security low security

Institution Name

Serial Number

Grid Header

lists the steps and dispense volumes of the selected protocol template for the current staining run. Each column represents one step of the protocol template.

Command Buttons:

Slide Info – enter or

edit individual slide information. Protocol Template – select or create a template to apply to the current staining run. Next – displays the Slide Layout Map: Program Slides screen. Print – prints slide and reagent labels, main grid, IHC report, reagent list, or Run logs. Exit – returns to Main Menu without saving. Help – information on grid features.

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Section 4

Entering Slide Information

Slides can be programmed from the Programming Grid either through menu bar for quick programming of total slide count, or through the Slide Info button for entering specific slide identification information.

Adding Slides

The Slides option expedites programming by reducing the amount of information required to program a staining run.

1. Select Slides on the menu bar. This function is used to bypass the Slide Information

screen when programming a staining run.

2. Enter the desired number of slides in the text box or use the scroll bar to select the

desired number of slides and click OK.

Note: Once a run has been set up using this function, slide, tissue, or doctor information cannot be added to the run. Slide Identification, Case #, Block ID, Tissue, or Doctor Name will not be associated/displayed in the Programming Grid when the Slides function is used. If the Slide Info button is selected the Slide Count text box appears rather than the Slide Information screen.

Slides on the

The Slide Info Button

The Slide Info button allows you to enter specific information for individual slides. The fields displayed are determined during initialization.

1. Select the Program button on the Main Menu screen.

2. Select the Slide Info button on the Programming Grid and enter slide information.

Note: An entry is required to proceed from one field to the next. Each field requires at least one key stroke. Type the specified information or press the SPACE BAR before moving to the next field. The Slide/case field requires entry of the actual number of slides.

3. Select the Finish Entry button or press ENTER until the Finish Entry button is

highlighted to complete the slide identification entry process. The Programming Grid appears.

Autostainer Handbook

Deleting Slides

Deleting Slides With the Slides Function

When using the deleted if there is at least one entirely blank (unprogrammed) row. Select the menu item, adjust the number of slides, and click OK.

Note: If all rows are programmed, the slide must be deleted from the Programming Grid (see Deleting a Specific Slide, later in this section).

Deleting Slide ID Information Using the Slide Info Button

Deleting Slide Identification – deletes all slides associated with the selected slide

identification. Deleting a Case Number – deletes all slides associated with the selected slide identification’s case. Deleting a Block ID – deletes all Block IDs associated with the selected case number.

1. Select the Slide Info button from the Programming Grid.

2. Press the DOWN ARROW key until the desired slide identification is highlighted in

the Slide ID field and press ENTER. The cursor moves to the Case # field. Select additional criteria if needed.

3. Select the Delete button. A dialog box asks if you want to delete the selected slide

identification.

4. Select the Yes button. Select the Finish Entry button to save changes.

Deleting a Specific Slide This method deletes one slide at a time.

1. On the Programming Grid left-click the slide number on the row representing the

slide. A dialog box asks if you want to delete the selected slide.

2. Select the Yes button and the selected slide is deleted from the current program.

Note: If slide labels have been printed, the reagent names are replaced with blank tiles in the deleted slide row.

Slides option on the menu bar to program a staining run, a slide can be

Slides

Rearranging slide positions on the Programming Grid

1. On the Programming Grid right-click the slide number on the row representing the

slide.

2. Move the mouse pointer to the position where the slide is to be inserted and left-

click. The slide information is inserted in the new position.

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Section 5

Designing a Protocol

Protocol templates allow you to enter the series of steps required to stain a slide and save the protocol for future use. Low level security does not allow creating or editing protocol templates.

Standard Protocol Elements

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.

Endogenous Enzyme Block Protein Block Primary Antibody Pretreatment Secondary Reagent (e.g., biotinylated secondary antibodies or link) Tertiary Reagent (e.g., enzyme-labeled streptavidin or label) Labeled Polymer (e.g., used in the EnVision visualization systems) Substrate-Batch Substrate Auxiliary (e.g., counterstain)

Autostainer Handbook

Additional Protocol Elements

Rinse Buffer

A buffer rinse step consists of a buffer wash and air blow cycle and should be programmed between each protocol template step. Each buffer rinse is displayed as a blue droplet in the programming grid with a droplet icon in each programmed row. The buffer icon appears on the Programming Grid printout as a filled-in droplet.

Rinse Water

A water rinse step consists of a water wash and air blow cycle. Each water rinse is displayed as a white droplet in the programming grid with a droplet icon in each programmed row. The water icon appears on the Programming Grid printout as a white droplet.

Standard Rinse Replacement

The standard rinse cycle can be replaced by a blow only step (reagent is blown off the slide but no buffer is applied) by right-clicking on the droplet icon in the programming grid. The droplet icon is replaced by a wind symbol.

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 groups 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 (non-substrate) are completed. An audible signal indicates when the substrate should be loaded and the screen displays where it should be placed (typically in position D8 of reagent rack a).

Note: The Substrate-batch step can only be used once in a Protocol Template.

Switch (*Switch)

This step programs the switching of waste disposal from one container to another for the separation of hazardous from non-hazardous waste. The Switch step is displayed in the Programming Grid as a skull-and-crossbones icon in each programmed row. The Switch step can be used multiple times in any Protocol Template. When programming successive switch steps, the skull-and-crossbones icon alternates with a flower icon to indicate switching from hazardous to nonhazardous waste.

Note: The switch step must be preceded by a rinse step in the protocol template. The first switch step in any protocol template is a switch to hazardous waste.

Creating and Editing a Protocol Template

Note: Template names must contain eight or fewer alpha-numeric characters, and no

symbols or spaces should be used.

1. Select the Protocol Template button on the Programming Grid. The Protocol

Template Design screen appears with the default protocol template displayed in the Protocol Outline column.

2. Select the New Template button. The default protocol template is replaced by the

default first step (rinse buffer) of the new template.

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Note: If No Pre-rinse is selected at the Options screen, the first rinse buffer is not displayed (See Initializing, Section 3).

3. Build the new protocol template from the Protocol Elements list. Click on a reagent

in the list, the item is copied to the Protocol Outline list and sequentially numbered.

Note: If Add Rinse to Protocol Steps is selected at the Options screen, rinses are automatically added after each step, except following a substrate step.

4. To delete a step, select it in the Protocol Outline list. The name is outlined. Select

the Delete Step button.

5. Additional steps can be inserted in the Protocol Outline by highlighting a step and

clicking on the desired step from Protocol Elements. The selected step will be inserted above the highlighted area in the Protocol Outline.

Note: The Autostainer can accommodate protocol templates containing as many as 35 steps (including rinse steps between reagent incubations).

6. Protocol templates can be used with or without saving. To save a newly created

template select the Save button on the Protocol Template Design screen. The

Save Template window appears with the cursor in the File Name box.

Deleting a Protocol Template

1. Select the Protocol Template button on the Programming Grid.

2. Select the Get Template button and click on the file name of the desired template.

3. Click the Delete Template button. A message asks if you want to delete the selected

protocol template. Click the Yes button.

Note: If the default template is deleted, a new template must be selected from the Get Template window and saved as the default template.

Using a New Protocol Template Without Saving

1. After a Protocol Template has been created or modified, click the Use Template

button in the Protocol Template Design screen. A dialog box asking “Save on disk now?” is displayed.

2. Select No and the Programming Grid appears with the new protocol template in the

header row.

Note: If a staining run had already been programmed and the new protocol template omits previously programmed steps, a warning message is displayed stating that programmed steps will be discarded if the new protocol template is applied.

Using a Saved Protocol Template

1. Select the Protocol Template button in the Programming Grid.

2. Select the Get Template button, select the template from the choices in the list, and

click OK.

Autostainer Handbook

3. Click the Use Template button. The Programming Grid appears with the selected

protocol template displayed in the grid header.

Using a Saved Protocol Template for a Specific Auto Program

Auto Programming is a feature for making slide programming more efficient (see Programming a Staining Run, Section 6).

1. Select the Protocol Template button in the Programming Grid.

2. Select the Get Template button.

3. Select the Auto File check box.

4. Select the desired Auto Program by clicking on it and click OK.

5. Click the Use Template button in the Protocol Template Design screen. The

Programming Grid appears with the selected protocol template displayed in the grid header.

Selecting Reagent Dispense Volume for All Steps in a Protocol

1. 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.

2. Select the raised diamond next to the appropriate volume and click OK. The selected

volume is applied to all steps in the protocol template.

3. Click the Use Template button. A message asks “Save on Disk now”. Select the Yes

button, enter a template name and click OK. Select the No button to use the template without saving the change. The newly selected dispense volume is displayed in the grid header of the Programming Grid.

Selecting Reagent Dispense Volume for a Specific Step in a Protocol

1. Select a protocol template step in the Protocol Outline list to assign a new reagent

dispense volume.

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. Select the desired reagent dispense volume by clicking on the raised diamond next

to the appropriate volume and click OK. The selected volume is applied to the specific step in the Protocol Template.

4. Click the Use Template button. A message asks “Save on Disk now”. Select the Yes

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Section 6

Programming a Staining Run

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 specific reagents for a staining run.

Each row represents one slide and its information.

Reagents that are available for the highlighted step.

Defining Detection Reagents

Before you can program specific reagents for slides, you must enter slide information and select a protocol template as described in Section 4 and Section 5. The Programming Grid is displayed listing the slides identified during Slide Count or Slide Information entry. The first programmable field is highlighted and a list of available reagents for the step is displayed.

1. Select a reagent using the UP and DOWN arrow keys and insert the reagent by

pressing ENTER. A message asks, “Assign to the following unprogrammed slides?”

2. 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 reagent is assigned to only the highlighted step.

3. After all reagents are programmed, click the Next button. The system checks the grid

for missing rinses. If a rinse is missing between two reagents, a message asks if you want to continue.

Autostainer Handbook

4. Select Yes and, if the program is new, a dialog box prompts “Save program on

disk?” Otherwise, the Slide Layout Map screen appears. Select Back to Grid to return to the Programming Grid and the missing rinse appears in the grid.

Defining Primary Antibodies

Each primary antibody reagent is programmed individually and will appear in the Programming Grid with its programmed pretreatment (see Reagent Management, Section 7). Primary antibodies and/or pretreatments can be manual steps and are noted on the Programming Grid and printouts. If a reagent is a manual step (i.e., overnight incubation, target retrieval, etc.), then it is performed before the slides are loaded.

Note: A primary antibody’s pretreatment automatically appears with the primary antibody only if the protocol template contains a pretreatment protocol element (see Designing a Protocol, Section 5).

1. Enter slide information, select a protocol template, and add necessary detection

reagents. When the first programmable Primary Antibody is highlighted, a list of available Primary Antibodies for the step is displayed.

2. Select the desired antibody from the list using the UP and DOWN arrow keys on the

keyboard and press ENTER.

Note: To make a pretreatment a manual step, right-click on a pretreatment. The list of available pretreatments is displayed. Select the desired pretreatment from the list.

3. Universal negative controls and previously assigned and saved matched negative

controls are listed at the end of the primary antibody list and are accessed on the keyboard by pressing the SHIFT key and typing “_”. Once the correct negative control is selected, press ENTER and the selected negative reagent and pretreatment will appear in the Programming Grid.

4. Select the Next button, the system checks the grid for missing rinses. If a rinse is

missing between two reagents, a message asks if you want to continue.

5. Select Yes and, if the program is new, a dialog box prompts “Save program on

disk?” Otherwise, the Slide Layout Map screen appears. Select No to return to the

Programming Grid and the missing rinse appears on the screen.

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 are added as new slides and contain the matching detection reagents of the test specimen.

20 Autostainer Handbook

Adding Positive and Negative Control Reagents

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. Click OK or the Finish Entry button.

2. Fill in Programming Grid columns with specific reagents for one slide. After the

primary antibody is applied, Neg.Ctl and Pos.Ctl buttons display in the menu bar. Continue entering the detection reagents.

3. To insert a matching positive control, click the Pos.Ctl button. One slide will be

added to the Programming Grid with reagents that match the last programmed slide. In the slide number column, PC appears signifying a positive control reagent.

Negative Controls

Two options are available for Negative Controls:

 Unique negative controls for each primary antibody.

 Universal Negative Controls that use a generic negative control for all antibodies of a

certain species. The Autostainer uses two different types of negative control, N series (mouse and rabbit) and NP series (mouse and rabbit). The Neg.Ctl button automatically assigns the appropriate universal negative control such as, UNC for N series antibodies or UNC+ for NP series antibodies. The NP series is used with higher sensitivity kits like EnVision+, LSAB+.

Dako Universal Negative Controls (UNC) are available for ready-to-use antibodies. If the Universal Negative Control option is chosen, all matching negative controls designated as “_NC” (i.e., NCVimentin) on the primary antibody list must be deleted from the reagent list.

Note: If you require assistance deleting the matched negative controls, please contact your Dako representative.

Assigning Negative Control Type to Antibodies

1. From the Programming Grid select Edit Lists from the menu bar.

2. Select the Primary Antibody category from the reagents list.

3. Select a previously defined antibody or add a new antibody to the list.

4. Enter the correct compatibility code for the antibody (see Compatibility Check in

Section 7).

5. Select UNC or UNC plus from the Neg.Ctl Type field to assign a Universal Negative

Control.

Using the Negative Control Tool To insert a Universal Negative control, click the Neg.Ctl button in the Programming

Grid.

Note: The Neg.Ctl tool selects one of the four Universal Negative Controls based on the

compatibility and negative control type defined for the primary antibody. If the primary

Autostainer Handbook

antibody does not have either a compatibility or negative control type defined, an error appears stating that there is no matching negative control reagent.

Note: If you do not wish to use this feature noted above, then continue programming as usual. This feature is not active when using Auto Program.

Defining Specific Rinses

A rinse is automatically added following every programmed reagent (except for Protein Block, where a blow step is added) when assigned as described above. The type of rinse is selected in the Protocol Template screen (see Designing a Protocol, Section 5).

The type of rinse can also be changed from the Programming Grid.

Click an activated tile in a rinse column. Click again to change the rinse to a different selection: rinse buffer, rinse (DI) water, blow, or none.

Note: Rinses that precede a switch cannot be changed from the Programming Grid. The rinse at the end of a run cannot be changed into a blow step.

Auto Programming

Auto Programming is a feature to make slide programming more efficient. This function

is used to save all or any portion, of a protocol to be used in future programming. An Auto Program can consist of one or more rows of programming. Low level security does not allow access to creating and saving Auto Programs.

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. Click OK or the Finish Entry button.

2. Fill in the Programming Grid columns with specific reagents, except primary

antibody, for one of the programmed slides.

Note: This creates an Auto Program for the detection reagents only. To create an Auto Program for a protocol that includes the primary antibody, enter specific primary antibody and detection reagents in the programming grid columns.

3. Select Auto from the menu bar on the Programming Grid.

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. Click and

drag to select additional rows. Each selected row is highlighted. Release the mouse button and the Save Auto Program window appears.

6. Enter a file name for the saved protocol. The Auto Program file name must contain

eight or fewer alpha-numeric characters, and no symbols or spaces can be used. Select the OK button.

7. Select Setup from the menu bar. Setup changes to Auto and exits the Auto

Program function.

Note: To use a saved Auto Program, the template you are programming must contain the same protocol elements, listed in the same order, as the saved Auto Program. To find the correct protocol template, use the Auto file function. The dispense volumes do not have to be the same.

22 Autostainer Handbook

Deleting an Auto Programming Item

1. Select Auto from the menu bar on the Programming Grid.

2. Select Setup from the Auto menu. The word Auto changes to Setup.

3. Select any activated tile in the Programming Grid to display the Save Auto

Program window.

4. Click the Auto Program to be deleted.

5. Click the Delete Program button.

6. Click the Yes button to delete the program.

Using an Auto Programming Item

1. Select Auto from the menu bar in the Programming Grid.

2. Select Program from the Auto menu.

3. Select a tile in the row where the Auto Program is to be inserted. A list displays the

saved Auto Program files that can be assigned for this slide.

4. Select the desired Auto Program file name to insert the program.

5. If the Auto Program does not contain a primary antibody, a message prompts

“Assign to all following slides.”

6. Click Yes to assign the Auto Program to all unprogrammed slides. Click No to assign

the Auto Program to only this slide.

Note: Skip is the first item in the Auto Program list. Select this option when you want to leave a slide unprogrammed.

7. Select Program.

8. Click the Next button. The system checks the grid for missing rinses. If a rinse is

missing between two reagents, a message asks if you want to continue.

9. Select Yes and, if the program is new, a dialog box prompts “Save program on

disk?” Otherwise, the Slide Layout Map screen appears. Select No to return to the

Programming Grid and the missing rinse appears in the grid.

Copy/Paste

1. The Copy/Paste feature copies reagents from one slide to another during slide

programming.

2. Select Copy from the menu bar on the Programming Grid. The word Copy

changes to Select.

3. Use the mouse pointer to select the steps to be copied by clicking and dragging

across the desired tiles. Once the tiles or rows have been highlighted, the word Select will change to Paste.

4. Click in the area where you wish to insert the copied selection.

Note: Copied rows can only be pasted under the same heading and cannot be pasted over areas that are currently highlighted.

5. Select Paste on the menu bar to deactivate the Copy function.

Autostainer Handbook

Editing a Protocol Step

The Autostainer software provides multiple methods for editing the reagents in a defined staining run from the Programming Grid.

Changing a reagent for a step:

1. Click the reagent step you wish to change.

2. Select the desired reagent from the list. The new reagent is displayed for the step.

Replacing a reagent with skip step (no reagent added to slide):

1. Click the reagent step you wish to change.

2. Select the word None.

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 the appropriate step.

2. Select Edit Slide from the reagent list. The Edit Individual Slide Reagent window

displays the information associated with the reagent.

Note: Lot and date cannot be edited here and are grayed out. The reagent long name, short name, compatibility, and incubation time can be edited.

3. Add a new reagent by entering the appropriate information.

4. Select the OK button. The new reagent is added for the current staining run only and

will not be added to the available list of reagents for this step.

Note: Do not use symbols when entering reagent information. Doing so may cause errors during the run.

Printing Options

Autostainer allows you to print slide and reagent labels as well as reports that include information regarding programs, runs, and stored reagents.

Slide Labels

1. Select the Print button at the bottom of the Programming Grid screen. The Which

Report screen displays seven options.

2. Select the Slide Labels button.

3. To print all slide labels for the program, select the All Labels button. The slide label

format is determined by the Design Slide Label screen that is accessed from the Initialize screen (see Initializing, Section 3).

4. To print a range of labels, type the range of labels with a hyphen between the

starting and ending numbers in the range (i.e., 5-12). To print individual labels, type the label number with commas between each number (i.e., 1,14,22). Both formats

can be used to print a combination of a range of labels with individual labels (i.e., 512,14,22). Do not use a space between numbers.

24 Autostainer Handbook

Reagent Labels

Printing reagent labels from a programmed run:

1. Program or retrieve a previously saved run on the Programming Grid, and select

the Print button.

2. The Which Report screen appears, click the Reagent Labels button.

3. To print all reagent labels, select the Print All button.

4. To print specific reagents, click the desired reagents and select the OK button.

Printing reagent labels not associated with a specific program:

1. From the Programming Grid select the Print button.

2. The Which Report screen appears, click the Reagent Labels button.

3. The Print Reagent Labels screen appears.

4. Select the Other Reagents button.

5. To print all reagent labels, select the Print All button.

6. To print specific reagents, click the desired reagents and select the OK button.

Reports

Program Grid Report

IHC Report Groups all slides by case number and includes slide numbers, primary

Run Log Report

1. From the Programming Grid, select the Print button.

2. Select the Program Grid, IHC, or Run Log button.

Prints the Programming Grid including all slide information, protocol headings, specific reagents, institution information, user, date printed, number of slide ID’s, number of case numbers, and the Autostainer serial number.

antibody information, and name and incubation time for all reagents used. Prints each step performed during a run.

Viewing Programmed Slides

When reagent programming is complete, the first Slide Layout Map appears. The Slide Layout Map screen is a representation of all 48 slides as they would be viewed facing

the front of the Autostainer. The dispense volume and drop zone(s) for each slide can be changed on this screen. The location for reagent dispense is displayed by a yellow highlight.

Autostainer Handbook

Assigning Reagent Dispense Locations

The non-frosted area of each slide is equally divided into three horizontal sections – or dispense locations. Each slide can have one to three dispense locations, and the maximum dispense volume of a slide cannot exceed 800 µL. The default dispense location is programmed during initialization. Modifications to the dispense volume or dispense location of a slide in the Slide Layout Map: Program Slides will change the text color of that slide from black to red in the Slide Layout Map.

Printing a Slide Layout Map

The Slide Layout Map provides a record of reagent dispense volume and dispense location. It is useful for ensuring that slides are loaded correctly.

1. Select the Next button on the Reagent Layout Map (see Section 8).

2. Select the Print button.

Selecting a new reagent-dispense location for all slides:

1. Select the All Slides tool in the upper left corner of the Slide Layout Map: Program

Slides.

2. Click the desired location on the slide to select a new reagent dispense location.

3. To assign multiple dispense locations on the All Slides tool, press and hold the

CTRL key while clicking on the desired dispense locations.

4. Click the dispense volume. A message displays the dispense volume choices.

5. Select the diamond next to the desired dispense volume.

6. Select the OK button or press ENTER.

7. The Slide Dispense Tool enables the user to make these changes for a specific

slide. The Slide Dispense Tool button tool functions identically to the All Slides tool except the user is able to manipulate drop zone and reagent application amounts to only the selected slide.

26 Autostainer Handbook

Section 7

Reagent Management

Reagent lists are created and modified using the commands on the Edit Lists menu on the Programming Grid.

Adding Detection Reagents (Secondary, Tertiary, Substrate, etc.)

1. From the Programming Grid select Edit Lists from the menu bar.

2. Select the reagent category from the list.

3. Enter the full name of the reagent you want to add to the current list and press

ENTER. The software allows the entry of 33 alpha-numeric characters for detection reagents. No symbols can be used in the reagent Long Name field.

4. Enter an abbreviated name for the reagent in the Short Name field and press

ENTER. The abbreviated name cannot exceed 10 alpha-numeric characters and should be unique to distinguish it from other reagents. No symbols can be used in the reagent short name. The cursor moves to the Compatibility field.

5. Enter the correct compatibility code and press ENTER. Assigning compatibility

information to reagents is optional (See Compatibility Check at the end of this section).

6. Enter the reagent lot number and press ENTER. The lot number cannot exceed 8

alpha-numeric characters. Lot number information is optional.

7. Enter the reagent expiration date and press ENTER. The reagent expiration date is

optional. Use MM/YY format.

Note: All expiration dates are limited to a maximum of 10 years from the date of entry.

8. Enter the reagent incubation time and press ENTER. Follow steps 3-8 until all

reagents have been added.

Note: A minimum of two minutes incubation time is required for proper operating conditions.

9. Select the OK button to add the new reagent(s) to the Reagent List.

Autostainer Handbook

Adding Primary Antibodies

A primary antibody can be added to the reagent list with a related pretreatment reagent. This facilitates automatic selection of the pretreatment whenever a primary antibody is selected from the Programming Grid.

1. Follow the same steps as in the Adding Detection Reagents section.

Note: To add a matching negative control use exactly the same name as the positive control and add the prefix “_NC” to the beginning of the reagent name.

2. If the primary antibody is a step that is preformed off the Autostainer (i.e., overnight

incubation) or requires heating, check the Manual Step check box by pressing the SPACE BAR.

3. Enter the correct compatibility code and press ENTER. Assigning compatibility

information to reagents is optional (see Compatibility Check at the end of this section).

4. Select the pretreatment from the box by using the UP and DOWN arrows, or type a

new pretreatment name, and press ENTER

Note: If Manual Step is selected, the incubation time will be “0”. Do not change this time.

Editing and Deleting Reagent Lists

1. From the Programming Grid select Edit Lists on the menu bar.

2. Select the reagent category from the list.

3. Edit fields by highlighting the information and typing over it. When changing the

reagent name, the software saves the change as a new reagent.

4. To delete, click the Delete button.

5. Select the OK button.

Updating a Reagent Lot Number and Expiration Date

1. From the Programming Grid select Edit Lists on the menu bar.

2. Select the reagent category from the list, and choose a specific reagent.

3. Highlight the current lot number displayed in the lot box. Type the new lot number

and the Change button appears.

4. Highlight the date displayed in the box and enter the new expiration date.

5. Click the Change button – all fields clear.

6. Select the OK button until a dialog box prompts “Save changes on disk?”

7. Select Yes to save the new lot number and expiration date. Select the No button and

the reagent is not edited.

28 Autostainer Handbook

Compatibility Check

A compatibility check is included 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 for the species reactivity compatibility of the primary antibody and the secondary reagent. The second addresses the compatibility rules dictated by the enzyme used in the detection system. The following tables summarize the compatibility rules and codes assigned to each reagent type.

Every reagent in the reagent categories list (or 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 antibodies can be marked with compatibility code AB. The software verifies that these compatibility rules are not violated during the programming of a staining run. If an incompatible reagent is assigned to a tile in the programming grid, a message displays the two reagents that are incompatible.

Species Reactivity

Reagent Type

(Protocol Element)

Primary Antibody Monoclonal primary antibodies raised in

mouse (e.g., mouse anti-human)

Primary Antibody Polyclonal primary antibodies raised in

rabbit (e.g., rabbit anti-human)

Secondary Reagent Secondary reagents (antibodies)

compatible with monoclonal antibodies raised in mouse (e.g., biotinylated antimouse IgG)

Secondary Reagent Secondary reagents (antibodies) reacting

with polyclonal antibodies raised in rabbit (e.g., biotinylated anti-rabbit IgG)

Secondary Reagent Secondary reagents (antibodies) reacting

with both monoclonal and polyclonal antibodies raised in mouse or rabbit, respectively

Enzyme System Compatibility

Reagent Type

(Protocol Element)

Endogenous Enzyme Block, HRP-compatible X Endogenous Enzyme Block, AP-compatible Y Tertiary Reagent HRP-labeled streptavidin X Tertiary Reagent AP-labeled streptavidin Y Substrate HRP-compatible substrates (e.g., DAB, AEC) X Substrate AP-compatible substrates (e.g., Fast Red,

New Fuchsin)

Description Compatibility

Code

Description Compatibility

Code

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30 Autostainer Handbook

Section 8

Loading Reagents

After programming is complete, the Autostainer calculates the time required for the staining run, the amount of wash buffer or deionized water required, and the most efficient placement for the reagent vials.

Reagent Layout Map Screen

The Reagent Layout Map documents the rack positions of the reagents including the fill volume needed for completing the staining run. It displays a maximum of 32 reagent vials per rack. The abbreviations for the reagents are displayed in their appropriate alpha-numeric rack positions.

Each reagent position is color coded to correspond to Dako reagent colors and the colors from the Programming Grid. If more than 32 reagent vials are required, a Second Rack button becomes available.

1. Select the Next button in the Slide Layout Map: Program Slides, a message asks,

“Save program on disk?”

2. Select No and the Reagent Layout Map appears.

3. Load the reagent vials according to the reagent map. The reagent volume displayed

includes a dead volume of at least 200 µL.

4. Make sure that enough reagent is in the vials to complete the programmed run and

that the reagent rack is properly seated in its correct location.

Autostainer Handbook

5. Check that the reagent vials have no bubbles on top of the solution. Remove any

bubbles.

Missing Reagent Notice

The Autostainer displays a prompt when an insufficient volume of reagent is detected. You can pause the system to add more reagent to the specified vial. If sufficient volume of a reagent is not available, and you do not respond to the alarm, the Autostainer continues the run and provides a list of skipped or partially applied slides on the Run Log.

To program the system to check for sufficient volumes at the start of the run, select the Check Volumes option on the Set Start Time screen (see Starting a Run, Section 9).

32 Autostainer Handbook

Section 9

Loading Slides

After programming is complete and the reagents are loaded, you are ready to load the slides on the slide racks. Print the Slide Layout Map: Load Slides screen by selecting the Print button at the bottom of the screen. This map provides a guide for loading slides and can be saved as part of your staining record.

Note: Soak slides in a buffer solution containing 0.05% Tween-20 for a minimum of five minutes prior to loading the Autostainer.

Load Slides

Autostainer has four slide racks that hold up to twelve slides each. Slide racks 1-4 have slide positions 1-12, 13-24, 25-36, and 37-48, respectively. The first rack is located toward the front of the instrument. Racks two, three, and four are located behind the first.

1. Load slides into the racks by inserting the label end of the slide (with the specimen

facing up) into a slide clip. The rack may be positioned in the angular setting to facilitate slide loading.

2. Load each slide, in numerical order from left to right, into the slide rack according to

the Slide Layout Map: Load Slides printout.

3. Once all of the slides have been loaded into their correct slide positions and the slide

racks are properly positioned on their locating pins, gently pour buffer over the slides to keep them from drying out. For best results, pour buffer over the slide label instead of directly over the specimen. The buffer will then flow over the specimen without washing it off.

Note: When the slides have been loaded, put the rack back to its horizontal position.

Autostainer Handbook

34 Autostainer Handbook

Section 10

Starting a Staining Run

Preparing the System for a Staining Run

After the specimen-containing slides and reagent vials are loaded, perform the following checklist:

Slides

• Deparaffinized and rehydrated, soaking in buffer containing 0.05% Tween 20 using a squirt bottle to prevent slides from drying while loading additional slides.

• Seated securely in slide rack.

Racks

• Properly seated in the Autostainer.

Reagents

• At room temperature.

• Placed at proper locations in the reagent racks.

• Contain no air bubbles.

• Contain at least the reagent volume in each vial as specified in the Reagent

Layout Map.

Buffer

• Contains at least the volume of buffer indicated in the Reagent Layout Map plus any additional volume listed for the completion of a delayed run.

Water

• Contains at least the volume of water indicated in the Reagent Layout Map.

Waste Containers

• Have enough room to accommodate the volume of buffer and water used during the staining run.

Quick Start

1. Turn on the computer system. Double-click the Autostainer icon.

2. Enter User Name and password and press ENTER.

3. Select the Program button on the main menu.

4. Select Slides from the menu bar. (To enter information for a specific slide see

Section 4.)

5. Enter the slide count for the staining run and click OK.

Note: Skip steps six and seven if you are using the default template.

Autostainer Handbook

6. Select the Protocol Template button and select a protocol template for the run.

7. Select the Use Template button. The Programming Grid appears with the selected

protocol.

8. Select the desired volume and drop zones for either all slides, or on an individual side basis.

9. Assign specific antibodies and reagents to all slides and select the Next button. The Slide Layout Map: Program Slides screen appears.

10. Select the Next button and the Reagent Layout Map appears.

11. Load reagents using the Reagent Layout Map. Check water and buffer volume, and make sure you have enough space for waste.

12. Select the Next button. The Slide Layout Map: Load Slides screen appears.

13. Load slides using the Slide Layout Map: Load Slides screen.

14. Click the Next button.

15. The Set Start Time screen appears. The system calculates the run time and amount of buffer and DI water needed to complete the run.

16. Select the Prime Pump (Water) button, and click OK.

17. Select the Prime Pump (Buffer) button.

18. Set reagent levels to Check Volumes or Do Not Check Volumes.

19. Select the Start Run button.

Run Log Screen

During a staining run the Run Log screen is updated with each step performed.

• The New Program button is used to program a new run while the current run is

in process.

• The Review Program button is used to return to the Programming Grid to view

the run in progress.

• The Emergency Stop button can be used to stop a run at any time.

Shutting Down the Autostainer

1. Select the Sign Off button on the Main Menu.

2. A message prompts, “Quit using Autostainer?” Select the Yes button.

3. Shut down the computer.

36 Autostainer Handbook

Section 11

Maintenance and Troubleshooting

The Autostainer should be cleaned after 150 slides have been processed or after one week, whichever comes first. The instrument keeps track of the number of staining runs performed and displays a maintenance message listing the number of slides since the last cleaning cycle.

Note: Please contact your Dako representative regarding maintenance details.

Troubleshooting

Problem Possible Cause Course of Action

Autostainer will not turn on (top green LED not on). Message displays “Instrument Not On” when a run is started.

The Autostainer does not respond when you initiate a prime pump or a run (bottom LED not on). Message displays “Instrument Not On” when a run is started.

Autostainer suddenly stops during a run.

Buffer or DI Water does not flow out during a run or a prime.

Not enough buffer or DI

Bent or pinched tubing.

The Autostainer is unplugged at the power source.

The cable between the Autostainer and the computer is not connected properly.

The Autostainer will stop frequently during the run. This is due to set incubation times. Check the run log to verify.

Autostainer was not primed properly.

water in the appropriate reservoir.

Check the power cord connections and verify the surge protector is plugged in and turned on.

Shut down the system and turn the power off. Disconnect and reconnect the cable between the Autostainer and the computer.

Please use caution. Pins can be bent and cause damage to the instrument during operation.

No action is needed. The Autostainer will resume movement. Check Run Log screen.

Prime pump. Pump may need to be activated several times.

Fill the reservoir with the proper level of fluid. Then prime the pump.

Straighten out the tubing and prime the pump.

Autostainer Handbook

Problem Possible Cause Course of Action

Buffer or DI Water does not flow out during a run or a prime. (Cont.) Inlet filters are clogged and

Tubing not completely

Faulty buffer / DI water

All waste collects into one waste container. Waste tubing not placed in

Faulty waste pump.

Waste backs up in the sink. Clogged sink filter. Clean the filter when the

Reagent probe is not dispensing reagent.

Clogged reagent probe. Contact a Dako

Bubbles on the surface of

The yellow three-way

Z-head assembly is hitting slide racks during priming or during run.

Blow head/wash head is

Tubing is disconnected at the inlet fitting(s).

Check the tubing to verify that the fittings are secure. Check the inlet filter at the

are restricting fluid flow.

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. Ensure that buffer and DI

inserted into carboy.

water tubing is fully inserted into the proper carboys.

Contact a Dako pump. “Switch” is not programmed in the Protocol Template.

representative.

Program a “Switch” in the

Protocol Template.

Insure that all waste tubes the proper waste carboy.

lead to proper carboy.

Contact a Dako

representative.

instrument is not in use.

Using a small brush and

some water, remove the

material causing the

blockage. Insufficient reagent in vial. If the system detects

insufficient reagent to

complete the current step,

you have one minute to add

reagent, after which some

slides may be skipped.

representative.

Remove bubbles. the reagent in the vial.

Verify that the OFF position stopcock valve on the syringe has been moved.

of the valve is in the down

direction. The valve should

be in a “T” configuration. Arm is out of alignment. Emergency stop the run and

restart the run. Remove

steps already performed.

Recheck Reagent Layout

Map as this may cause vials

to be reassigned to new

locations.

Contact a Dako bent.

representative.

38 Autostainer Handbook

Problem Possible Cause Course of Action

The probe and wash/blow head are not going to the correct locations. Bent probe tip. Verify that all reagent vial caps have been removed

No Staining Insufficient reagent

Reagents vials were

Slides were not

Slides were loaded

Programming error. Check the Programming Grid to ensure that the

Slides drying out. Make sure adequate dispense volume that is

The text of the label does not begin along the left edge of the label and the text is getting cut off.

Error occurs during printing. Incorrect COM port

Incorrect printer is

The arm has been knocked out of calibration.

in vial.

not loaded in the correct locations in the reagent racks.

loaded in the correct locations.

with tissue side down.

The Seymour printer may need to be reset.

Labels or ribbon have run out.

selected.

Abort run and save run log. Reprogram the run and delete the steps from the Protocol Design screen that have already been completed. Restart run.

and slide racks are firmly locked in place in the down position. Contact a Dako representative. If the system detects insufficient reagent to complete the current step, you have one minute to add reagent, after which some slides may be skipped.

Check the Reagent Map to verify proper locations. Check Run Log for “Unable to Locate Reagent” Skipped Slides message.

Check the Load Slide printout to verify proper locations.

Orient slides properly.

staining run was programmed correctly.

suitable for desired incubation times is programmed. Insure that no extreme environmental conditions are contributing to the drying of the slides (i.e. direct sunlight, heating/air conditioning vents, hoods, etc.) 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 three seconds. The labels advance and the printer will print. Release the feed button. Press and release the feed button one more time to advance one label. Perform a test print. Replace ribbon in the printer and the printer will restart. Select the correct COM port from the Slide Label Design screen, see the Initializing section. Select the correct printer model from Slide Label Design screen, see the Initializing section.

Autostainer Handbook

Problem Possible Cause Course of Action

Slide number is not listed on label. Computer functions lock up during a run, but the Autostainer is still performing the run. User is unable to access many of the functions in the Initialize section of the program. Illegal or fatal exception errors.

EWP/DWP messages on Run Log.

EWL/PFR messages on Run Log. The computer turns on but “Safe Mode” is displayed on all four corners of the screen.

Monitor screen went blank (power LED is green). Monitor screen went blank (power LED is orange).

Label is not centered correctly on label. Windows error. Allow the Autostainer to finish the run. Reboot system

User has been set up as a “NonAuthorized” user.

Windows error. Shut down the system and turn the power off at the

Enable waste pump/disable waste pump.

Faulty waste removal/probe failed to retract. The computer has been improperly shut down.

Screen saver utility installed.

The cable between the monitor and the computer is not connected properly.

Reprint label if necessary and place the label 1/2 cm from the top and 1/8 cm from the side of the slide.

before the next run.

Contact a Dako representative.

surge protector. Wait 30 seconds and then power up the system. If this problem persists, contact a Dako representative. Waste is not being pumped out of the system within the expected time frame. If waste is still separating, contact a Dako Technical support for additional cleaning instructions. If waste is no longer separating, contact Dako for service. Contact a Dako representative.

Shut down the computer by using the Windows Start button located at the bottom left corner of the screen and selecting Shut Down. This button may be hidden requiring you to drag the mouse pointer off the bottom of the screen for it to appear. Always shut down the computer using this method. Perform Scan Disk through Windows Explorer/Accessories/System Tools/Scan Disk. For assistance, contact a Dako representative. Move the mouse. Current screen will reappear. Remove screen saver as this may interfere with the Autostainer operation.

Disconnect and reconnect the cable between the monitor and the computer.

40 Autostainer Handbook

Problem Possible Cause Course of Action

Words on your monitor screen are too dim. Image on the desktop is grainy and the color is incorrect.

Color depth has

The printer does not respond to print command. The parallel

The parallel

The brightness control is not set properly. Windows started in “Safe Mode.”

been adjusted too low.

The printer may be in Manual Feed mode.

cable between the printer and the computer is not connected properly.

cable is defective.

Adjust the brightness control.

Shut down the computer by using the Windows “Start” button located at the bottom left corner of the screen and selecting Shut Down. This button may be hidden requiring you to drag the mouse pointer off the bottom of the screen for it to appear. Always shut down the computer using this method. Perform Scan Disk through Windows Explorer/Accessories/System Tools/Scan Disk. For assistance, contact a Dako representative. At the Windows Desktop, use the mouse to right-click anywhere other than an icon, a drop-down menu displays. Select Properties. The Windows Display Properties window should appear. Select the Settings tab. In the Colors box verify that “High Color (16 bit)” is displayed. If it is not, select it and save changes. Briefly press and release the Front Panel Button on the printer.

Disconnect and reconnect the parallel cable between the printer and the computer.

Check the cable on another system with a print job that you know works, or connect another parallel cable.

Autostainer Handbook

42 Autostainer Handbook

Section 12

System Specifications

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. (63.5 kg) Electrical Requirements 120V 110/120V (±10%) 60Hz (±2Hz)

220V 220/240V (±10%) 50Hz (±2Hz) Normal Operating Temperature 18°C – 26°C (64°F – 79°F) Slide Rack Capacity 1 – 12 glass slides Total Slide Capacity 1 – 48 glass slides Reagent capacity 64 different reagents (15 mL/reagent vial) Reagent dispense volumes 100, 150, 200, 400 and 600 µL Reagent probe volume capacity 100 µL minimum, 1200 µL maximum

Computer Controller

Dako reserves the right to change the computer controller specifications at any time. Computer specifications may vary.

Monitor 15” color Printer Color InkJet printer Surge Protector 120 VAC, 15 Amps, 50/60 Hz

Software Specifications

Operating System Operating Logic Designed to calculate the most time-efficient sequence to complete a

Protocol Logic Flexible selection of steps including rinses. Maximum 35 protocol

Idle Rinse Buffer is applied to slides during a staining run to keep specimens

Rinse Default Settings

Incubation Time Recommended minimum time is two minutes.

Autostainer Handbook

Windows® 98, XP or equivalent.

programmed staining run.

steps (including rinse steps).

wet when no reaction is occurring. Buffer rinse every 30 minutes prior to the start of a run, when delayed start is selected. Buffer rinse every 30 minutes during a staining run. Water rinse every 60 minutes after completion of run.

Default settings for time and the amount of buffer dispensed can be changed by a Dako representative upon request.

44 Autostainer Handbook

Surface

Size Environment

Power Source

Section 13

Standard Configuration

Ensure that the area for the instrument is a solid, level surface that can support the weight of the system (approximately 240 lbs.) Minimum work area dimensions: 2.0 m W x 0.8 m D x 1.0 m. H The work area must have an ambient temperature between 18°C – 26°C (64°F – 79°F), not facing direct sunlight and not effected by a hood or vent. Dako recommends a dedicated power source to prevent interference from other instruments or equipment.

Z Head Assembly

Back

X Axis Mechanism

Reagent Probe Wash Station

Left

Right

Reagent Rack b

Reagent Rack a

Y Axis Mechanism

Front

Slide Rack

Glass Slide

Sink (Beneath slide racks)

Slide Rack Handle

Autostainer Handbook

Dako Autostainer User Manual

Specifications and Main Features

Frequently Asked Questions

User Manual

Dako Autostainer Handbook

Copyright © 2006 Dako, Inc. All rights reserved.

User Resources

Installation Procedure

Relocation Procedure

Scope

Disclaimers

Contact Information

Table of Contents

Warnings, Precautions and Limitations

System Overview

Software Overview

Sign In Screen

Main Menu

Initialize

Clean

Sign Off and Help

Reagent Tracking

Prime Pump

Programming Grid Overview

Entering Slide Information

Adding Slides

The Slide Info Button

Deleting Slides

Rearranging slide positions on the Programming Grid

Designing a Protocol

Standard Protocol Elements

Additional Protocol Elements

Creating and Editing a Protocol Template

Deleting a Protocol Template

Using a New Protocol Template Without Saving

Using a Saved Protocol Template

Using a Saved Protocol Template for a Specific Auto Program

Programming a Staining Run

Defining Detection Reagents

Defining Primary Antibodies

Assigning Positive and Negative Control Reagents

Defining Specific Rinses

Auto Programming

Editing a Protocol Step

Printing Options

Viewing Programmed Slides

Reagent Management

Adding Primary Antibodies

Editing and Deleting Reagent Lists

Updating a Reagent Lot Number and Expiration Date

Compatibility Check

Loading Reagents

Reagent Layout Map Screen

Missing Reagent Notice

Loading Slides

Load Slides

Starting a Staining Run

Preparing the System for a Staining Run

Quick Start

Shutting Down the Autostainer

Maintenance and Troubleshooting

Troubleshooting

System Specifications

Hardware Specifications

Software Specifications

Standard Configuration