Tutorial for MasterPlex® CT v1.0 © 2006 Hitachi Software Engineering America, Ltd. All Rights Reserved.
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In this tutorial, we will be highlighting 8 powerful features of the MasterPlex CT v1.0 software: 1) Probe Height Adjustment 2) Wizard Interface 3) Action List Manager 4) Three Step Acquisition 5) Kit Manager 6) Seamless Integration 7) Fail Safe Acquisition 8) Remote Monitor http://www.miraibio.
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Probe Height Adjustment Being able to adjust the probe height is crucial for any Luminex user as it is the cause for many known issues such as: Sample empty errors Slow event rates Air in the system Probes being stuck in the down position Actuator errors in the software Adjusting the probe height must also be performed after any of the following events: A new type of plate is being used Probe has been removed (ex.
Thumbwheel Bolt The initial Adjust Probe window gives detailed instructions on adjusting the probe height. To adjust the vertical height of the sample probe: 1. Remove the clear plastic shield that covers the sample probe area. 2. Press the Eject button and place a plate in the plate holder. 3. Place the appropriate alignment tool in the plate: For a standard plate with flat-bottom wells – stack two of the larger (5.
4. After you have placed the appropriate alignment tools into the well, press the Location button and specify the well where you have placed them. 5. Now loosen the bolt that holds the probe (refer to the picture in the Adjust Probe window) with the hex driver that is provided within the left panel of the Luminex system where the heater block is stored. 6. Click on the Probe Down button to lower the probe arm. The probe height will get adjusted by the spacers in the well. 7.
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Wizard Interface MasterPlex CT provides a set of wizard driven interfaces to help familiarize new users with the software. Whether you are new to MasterPlex CT or new to the Luminex 100 platform in general, wizards will guide you through every step of the way. There are a total of 8 wizards that will automate everything you will have to do from starting up the Luminex system to shutting it down. You can find the wizard launch buttons under the Home tab in the left task bar under Wizard & Automation.
Here is the breakdown and a brief summary of the wizards: The Startup Wizard walks you through the warm up and startup preparation sequence. The Calibration & Control Wizard walks you through using the calibration and control beads. The Region Activation Wizard guides you through selecting the bead regions used in the multiplex assay.
Please note that there are detailed instructions provided at each step. The next window checks to make sure that all the devices are connected properly. If there are any red flags, please check to make sure that all the connections are secured properly and then click the Connect button to try again. Once all the connections are marked by green checks, click on Next to proceed. The next screen gives a status of the laser warmup period. While the lasers are warming up, you can accomplish other tasks.
In the next step of the Startup Wizard, you can perform the Prime procedure. Priming removes air bubbles from the fluidics sub-system of the Luminex device. You can specify the number of cycles for priming by clicking on the Up or Down arrow under Number of Cycles. Once you have set the Number of Cycles, simply click on the Prime button and priming will automatically begin by the Luminex system. The status bar shows you the progress of the current action. Once the Prime is done, click Next to proceed.
The next step takes you through the Alcohol Flush, which removes air bubbles from the cuvette. In this step, you can specify the location of your reagent, in this case 70% isopropanol. To do this, click on the Location button, which in this case is currently labeled Reservoir. The plate window will pop up allowing you to specify the plate type and well location of your reagent. Specify the Number of Cycles for the Alcohol Flush and click on the Alcohol Flush button to execute the command.
You are now done with the Startup Wizard and your Luminex system is ready for use. After finishing one wizard, you will have the option to move on to the next wizard in the setup process. http://www.miraibio.
Action List Manager The Action List Manager lets you customize and automate routine procedures such as Startup, Maintenance, and Daily Shutdown. You can find the Action List launch button under the Home tab in the left task bar under Wizard & Automation Add Item button Remove Item button As an example, let’s create a Startup action list.
Next, we want to specify the number of times a step is to be repeated and the well location for the operation. Let’s change the Wash step so that it will perform two washes. To do this, simply double-click on the Cycle column next to the Wash step. Then click on the up-arrow to increase the number of Cycles to 2. The Wash step will now be performed twice in our Action List. Next, we want to specify the location of the well for an operation. Let’s say we are using the MasterPlex plate for our operation.
Lastly, we will name our newly created Action List to “Startup Procedure” in the Action Name form and press the Save button to save it. You should now see it saved in the Action List. To execute a previously saved Action List, just select the procedure from the Action List Manager and press the Start Action button on the bottom. A Progress Monitor will appear showing you the progress of each step of the Action List. http://www.miraibio.
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Three Step Acquisition MasterPlex CT v1.0 allows just three simple steps to run an acquisition: 1) What – Specify the regions you are trying to detect. 2) How – Set the way you want an acquisition to run such as gate settings, sample volume, time out values, output file and folder name. 3) Where – Specify where the samples are on the plate. Tell MasterPlex CT what you are trying to detect by activating bead regions.
Let’s begin with the Region settings. First click the Region Modify button to activate appropriate regions. The Bead Regions window should pop up. Next, activate regions by clicking on the check box to the right of the region number. Once a region is activated, it will be displayed with a red highlighted background. http://www.miraibio.
To define bead region names, click on the Properties tab. Next, double-click the default name under User defined name and edit it. You can also edit the Minimum and Maximum number of events to acquire for each region. Tips: - Leave the Region Map Type at the default 100 regions since it contains all of the bead regions that you can select from. - The copy and paste functions will work if you copy a column of values from a spreadsheet and paste it into the User Defined Names column.
Now, let’s enter our parameters that will determine how the machine will run by clicking the Setup Modify button. The Acquisition Setup window should pop up. In the Auto Stop tab, you can specify how you would like to stop counting the type of events. http://www.miraibio.
In the Setup tab, you can specify the Double Discriminator gate values, the direction of which the acquisition will be carried out, and the output file settings. The last thing to do before starting the acquisition is telling MasterPlex CT where your samples are located in your 96-well plate. To do this, click on the Sample Modify button to open the Sample Manager window. http://www.miraibio.
To mark your sample wells, you can simply drag and click to highlight wells or hold down the CTRL key to select discontiguous wells. After selecting your samples, press the Mark As Sample button. You can now edit your sample names under the Sample Editor tab. Please note that the copy and paste features are also available here. You can copy a column of names from a spreadsheet and paste them directly into the Sample Names column. http://www.miraibio.
Congratulations! You have now completed the 3 steps needed to begin data acquisition. You have entered your: 1) bead region information 2) setup configuration 3) sample location You are now ready to begin data acquisition. Click the Acquisition button to run the plate. http://www.miraibio.
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Kit Manager MasterPlex CT has built-in capabilities to recognize predefined kit configurations so that you will not have to spend time creating them. Kit configurations contain instructions on running the acquisition as well as instructions for analyzing the results. Many kit configurations are available right from the MiraiBio website.
You can use this Kit Manager to: Import new kit files View general information about a kit Duplicate a kit file to produce alternate parameters Delete kit files Export kit files for use on other computers To import our file, press the Import button and browse to the file you just downloaded and press the Open button. To view specific information about a kit file, select the file and press the Update Lot button. http://www.miraibio.
The Plate Map tab displays background, standard, and control sample locations. The Regions tab displays the analyte panel names. http://www.miraibio.
The Samples tab displays the information regarding the standards, negative controls and quality control samples. The Standards tab displays the preconfigured standard values eliminating perhaps the most time consuming step of creating an assay template. http://www.miraibio.
Click on the Cancel button to close the Lot Information window and then press the Apply button to use the kit configuration file. Next, you will be prompted to select your unknown sample wells. To do so, simply highlight the unknown sample wells and click the Add button. Alternatively, you can select discontiguous wells by holding down the CTRL button and left-clicking the unknown sample wells with your mouse. After adding your unknown sample wells, click on the Apply button.
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Seamless Integration MasterPlex CT seamlessly integrates with other MasterPlex analysis packages such as MasterPlex QT (quantitative analysis software for protein assays), and MasterPlex GT (genotyping analysis software). Once the acquisition is complete, results can be sent to the analysis application for immediate analysis. To begin analysis after an acquisition is complete, click the Analysis button and select the downstream analysis program you would like to transfer your data to. http://www.
The analysis program you have chosen automatically launches. If you have used a kit configuration file, all the template information and properties for the analysis program are sent with the data and the calculations are performed automatically. http://www.miraibio.
Select the data type pull menu and view the Concentration data. All the unknown concentrations for each analyte region will be displayed. http://www.miraibio.
To view the Standard Curve graph, click on the Standard Curve button. To view the x-axis in log scale, right-click on the graph and select Set XAxis to Log Scale. http://www.miraibio.
To view multiple standard curves in the same graph, hold down the CTRL key and select the regions you are interested in. http://www.miraibio.
MasterPlex CT has many different options to view data. For example, to view a graph in 3-D, click on the 3D button in the top icon menu. http://www.miraibio.
Fail Safe Acquisition With MasterPlex CT, your data acquisition need not be “all or nothing.” The Fail Safe Acquisition feature in MasterPlex CT allows you to recover from various common user errors. For example, if you run your assay and then discover that one of your bead regions was not correctly activated, you can activate the correct bead regions and then classify the data without the need to rerun the assay.
While the data is being reprocessed, the data is presented in the same view as if it was being acquired in real-time. You can view scatter plot and histogram information for the reprocessed data under the Sample Monitor tab. http://www.miraibio.
If you discover that you are not using optimal settings during the run, click the Stop button. Change the configuration settings and then press the Continue button. The previously collected data will be reclassified according to the new configuration settings. http://www.miraibio.
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Remote Monitor Free yourself from the instrument. MasterPlex CT includes a remote client program that allows you to monitor your Luminex instrument from another computer within the same local area network (LAN). Whether you are running a plate, waiting for the warm-up to complete or just want to know when the lasers are going offline, the remote monitor client will let you monitor the instrument from the comfort of your own desk.
In case of an emergency, the Luminex instrument can be remotely paused by just simply clicking on the Pause button. In the Plate tab, you can monitor sample acquisition well-bywell in real-time. http://www.miraibio.
In the Data tab, you can view MFI data as it is being acquired well-by-well in realtime. In the Count tab, you can view the real-time bead count in the form of bar graphs. http://www.miraibio.
In the Device tab, you can monitor the status of the Luminex instrument. In the above screenshot, the progress of a Prime operation is being monitored remotely in real-time. As you can see, this powerful MasterPlex CT feature allows you to walk away from the instrument during time-consuming operations. http://www.miraibio.
You can also contact MiraiBio with any questions you have at (510) 337-2000 (USA phone number) or by sending an email to support@miraibio.com. MiraiBio A division of Hitachi Software 1201 Harbor Bay Parkway Suite 150 Alameda, CA 94502 Telephone 1.800.624.6176 1.510.337.2000 Facsimile 1.510.337.2099 Trademark Acknowledgments MasterPlex is a trademark of Hitachi Software Engineering Co., Ltd. Luminex® is a registered trademark of the Luminex Corporation.
