Detailed interface description

The In-Solution Digestion: Single Plate App is designed to mimic common manual in‑solution digestion procedures for successively adding digestion reagents to samples and then incubating.

In this App, a 96-well sample plate is placed at Bravo deck location 4, and up to four 96‑well plates containing digestion reagents are placed at deck locations: 5, 6, 8 and 9. During the protocol, the AssayMAP Bravo Platform transfers each of the digestion reagents into the sample plate, one at a time, with optional heated incubation steps after each reagent addition.

Figure. In-Solution Digestion: Single Plate App interface

The App interface includes the following four setup sections (A–D) and a control panel.

A: Select Method

The Select Method section enables you to load previously saved digestion methods.

 

Control	Description
Browse for a Method	Enables you to locate and select a method file. All method files are stored in the following location: C:\VWorks Workspace\Methods\AM In Solution Digestion Single Plate v1.0
Load Values	Populates the interface with the application settings that are associated with the selected method file and updates the Bravo Deck Layout display with the labware selections at the appropriate deck locations.
Method Loaded	Displays the name of the loaded method.
Method Status	Displays general information and error messages during method setup and at runtime. For details, see Error messages.

The In-Solution Digestion: Single Plate App includes several pre-loaded digestion methods that cover the following generic digestion sequences:

•	For general in-solution digestion use cases:

Denature/Reduce -> Alkylate -> Dilute -> Digest (Protease Addition)

•	For use with the Protein Cleanup v1.0 App:

Denature/Reduce - > Alkylate - > Dilute

Digest -> Acidification

These pre-loaded methods provide common, easy-to-follow examples that you can use for initial testing purposes or as templates for creating custom digestion processes.

 

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B: Input Sample Settings

In this section, you specify the following important details about the Sample Plate at deck location 4:

 

Failure to properly lid or not lid the Sample Plate could cause a collision, which could result in equipment damage.

 

 

Proper monitoring of the total liquid volume in the Sample Plate is critical for optimal pipetting behavior.

 

 

Sample setting	Description	Range of values
Sample Plate Labware	Defines the type of labware that will be used for the Sample Plate. For approved labware options, see Labware.	–
Sample Plate Lidded	Specifies if the Sample Plate will have a lid during the run: • If the check box is selected, the Sample Plate must have a lid at runtime. • If the check box is cleared, the Sample Plate must not have a lid at runtime. Lid requirements will be displayed in the updated deck layout, just before runtime.	–
Starting Sample Volume (µL)	Defines the volume of sample that will be present in each well of the Sample Plate at the beginning of the run. Only include wells that contain liquid (do not average across the entire plate).	0–1000 µL/well
Number of Full Columns of Samples	Defines the number of full columns samples that will be digested during the run (rounded up to the nearest full column). This value is important for defining the amount of force that the Bravo 96AM Head head uses when picking up pipette tips.	1–12 columns

 

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C: Input Addition Step Settings

In this section, you specify the following settings for each of the four reagent addition steps:

 

Failure to properly lid or not lid the Sample Plate could cause a collision, which could result in equipment damage.

 

 

Setting	Description	Range of values
Addition Name	Gives a meaningful name to each of the addition steps. Select from the list of names, or type a custom name for the step. The pre-programmed options include NONE, Denaturation, Reduction, Alkylation, Internal Standard, Diluent, and Protease.	–
Reagent Deck Location	Displays the deck location of the reagent plate to be used for each reagent addition step. These values are read-only.	–
Addition Volume (µL)	Defines the volume of reagent that will be added to the Sample Plate for each addition step.	0–1000 µL
Mixing Cycles	Defines the number of aspirate-and-dispense cycles that will be used to mix the samples after the reagent has been added. The mixing volume is 75% of the cumulative Sample Plate volume, up to the maximum capacity of the pipetting method, where • Capacity for 250 µL Pipette tips = 140 µL • Capacity for bare AssayMAP Probes = 250 µL	0–10 cycles
Incubation Time (min)	Defines the time duration that the sample plate will incubate after a reagent has been added to it. A value of 0 indicates that no extended incubation will occur.	0–300 minutes
Incubation Temp (°C)	Defines the temperature (°C) at which the Sample Plate incubation will occur. The default value is OFF (room temperature), which indicates that the Peltier Thermal Station will not be activated for the incubation. Note: The temperature setting controls the temperature of the Peltier Thermal Station. The temperature in the wells may be less than the set temperature. Using a thermal plate insert helps to minimize this difference. For thermal plate inserts, see the Labware Reference Guide in the Literature Library page of the Protein Sample Prep Workbench.	OFF or 4–75 °C
Pause After Addition	Specifies whether to pause the run after completing the addition step, which would enable manual intervention, such as off-deck incubation or introducing fresh pipette tips. • If the check box is selected, a pause will occur. • If the check box is cleared, no pause will occur.	–
Labware Selection	Defines the type of labware that will be used for the reagent plate. For a list of approved labware options, see Labware.	–
Plate Lidded	Determines if the Reagent Plate will require a lid during the run. • If the check box is selected, the Reagent Plate must have a lid at runtime. • If the check box is cleared, the Reagent Plate must not have a lid at runtime. Lid requirements will be displayed in the updated deck layout, just before runtime.	–
Use Tips for Addition	Specifies whether to use pipette tips to conduct reagent addition and mixing. • If the check box is selected, the 250 µL pipette tips will be used. • If the check box is cleared, the bare AssayMAP probes will be used.	–
Number of Wash Cycles	Specifies the number of washes at the wash station as the last step of each reagent addition. The Use Tips for Addition setting determines whether pipette tips or AssayMAP syringe probes are washed.	0–10 cycles

 

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D: Update Deck Layout

After setting the properties in sections A to C, click Update Deck Layout with new Settings. The deck image displays the following for the selected settings:

•	Bravo deck location for each reagent

•	The labware types required for each reagent

•	Whether or not each labware requires a lid

 

Failure to update the deck to reflect the settings could result in incorrect labware being placed on the deck, which could cause a potential collision. Ensure that you update the deck immediately before every run to ensure that labware changes, including lids, are displayed during deck setup.

 

 

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Control panel

The control panel on the far right side of the application interface provides the following controls and indicators:

•	Status. Indicates which step of the run is currently being executed.

•	Cumulative Sample Volume (µL). Displays the current volume (µL) contained within the sample plate wells.

•	Run Protocol. Executes a run using the settings shown in the App interface. After this button is clicked and before any hardware motion occurs, a series of error checks are conducted to verify that the digestion procedure has been properly specified.

 

Error check	Description
Sample Plate Capacity	Compares the Initial Sample Volume input to the Sample Plate Labware selection to verify that it does not exceed the labware volume capacity.
Reagent Plate Capacity	Compares each Reagent Addition Volume input to its respective Labware Selection to ensure that it does not exceed the labware volume capacity.
Cumulative Volume	Calculates the cumulative digest volume (Initial Sample Volume + sum of Reagent Addition Volumes) to ensure that the final digest volume does not exceed the Sample Plate Labware volume capacity.
Sample Plate Pipette Tip Compatibility	If a deep-well plate type is selected for the Sample Plate Labware, each addition step is verified to ensure that the Use Pipette Tips for Addition? option is selected.
Reagent Plate Pipette Tip Compatibility	If a deep-well plate type is selected for any of the four reagent Labware Selections, the corresponding Use Pipette Tips for Addition? setting is verified to ensure that pipettes will be used for the transfer.
No Addition Steps	Checks to see that at least one reagent addition step has been specified by ensuring that not all of the Reagent Names settings have been set to NONE.

If an error is identified, an error message will display in the Method Status box.

•

Pause. Pauses the run after the operation that is in progress has completed. For example, if you click Pause while the AssayMAP Bravo Platform is in the middle of a dispense step, the system will pause after it has completely finished the dispense step in progress. You can resume the run after the pause.

To stop a run in an emergency, press the red emergency-stop button on the pendant. The Bravo motion stops immediately. You might not be able to resume a protocol after an emergency stop.

Note: Interrupting the light curtain causes the Bravo motion to stop immediately.

•	Restore Defaults. Clears the App interface of all settings and deck setup information.

•	Toggle Full Screen. Changes the display of the App interface to show the underlying VWorks developer interface. Only experienced users should use the Full Screen Off view.

•	App Library, Utility Library, and Workflow Library. Returns the display to the corresponding page in the workbench.

•	Save Method. Enables you to save new method files, and if necessary, overwrite existing methods.

 

Control	Description
Name	You may enter the name of a new method, or the name of an existing method that you want to overwrite with new settings.
Overwrite if Name Exists	To overwrite an existing method, select this check box. This is an insurance mechanism to prevent the unwanted overwriting of method files.
Save	Saves the method using the settings that are currently shown in the App interface.

When a new method is saved, a primary copy of the method file is saved in the following location:

C:\VWorks Workspace\Methods\AM In Solution Digestion Single Plate v1.0

The software also creates a backup copy when a new method is created or an existing method is overwritten. The backup copy enables recovery of a method file that has been unintentionally overwritten or lost by other means.

The software appends a date and time stamp to the file name of the backup copy. For example, if the method file name is Test Method 1, the backup copy file name would be as follows:

Test Method 1_Mar 3 2015 16_03_25

where March 3 2015 is the month, day, and year; and 16 is the hour, 03 is minutes, and 25 is seconds.

The backup method files are saved in the following location:

C:\VWorks Workspace\Workbench\Backups\AM In Solution Digestion Single Plate v1.0

 

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