Lum/nex ® Luminex 100™ IS Developer Guide to xMAP™ Technology Version 2.
© LUMINEX CORPORATION, 2003. All rights reserved. No part of this publication may be reproduced, transmitted, transcribed, or translated into any language or computer language, in any form or by any means without prior express, written consent of: LUMINEX CORPORATION 12212 Technology Boulevard Austin, Texas 78727-6115 U.S.A. Voice: (512) 219-8020 Fax: (512) 219-5195 Luminex 100™ IS Developer Guide to xMAP™ Technology Version 2.1 PN 89-00002-00-062 Rev.
Contents Luminex xMAP Technology 1 Luminex 100 IS xMAP Technology . . . . . . . . . . . . . . . . . . . . . . . . 1 Overview. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 About This Guide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 Performance Specifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Speed. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology Luminex 100 IS Developer Workbench Software . . . . . . . . . . . . 21 Developer Workbench Procedures . . . . . . . . . . . . . . . . . . . . . . . . 22 Create Products . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22 Definitions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22 Create New Product Dialog Box Buttons . . . . . . . . . . . . . . . . .
xMAP Technology Microsphere activation. . . . . . . . . . . . . . . . . . . . . . . . . . . . Coupling, blocking, and storage. . . . . . . . . . . . . . . . . . . . . Technical Notes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . One-Step Carbodiimide Coupling of Oligonucleotides to xMAP Carboxylated Microspheres . . . . . . . . . . . . . . . . . . . . . . . Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Equipment. . . . . . . . . . . .
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 iv xMAP Technology PN 89-00002-00-062 Rev.
L Luminex xMAP Technology Luminex 100 IS xMAP Technology The Luminex 100 IS uses software specially developed to increase efficiency in the clinical laboratory and research and development. It assumes that end users in the laboratory are running microtiter plates using xMAP-based assay kits developed for use with xMAP technology. Include in your kits to end users a template on floppy disk or CD with instructions. Overview Use the xMAP Technology software to develop templates for use on the Luminex IS.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.
xMAP Technology Capacity Luminex xMAP Technology The following specifications reflect minimum capacity values: • 10 GB hard drive • Store data for up to 240,000 test results • 1.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology General Guidelines Warning: Human/animal samples may contain biohazardous infectious agents. Where exposure (including aerosol) to potentially biohazardous materials exists, operators should follow appropriate biosafety procedures and use personal protective equipment, such as gloves, gowns, laboratory coats, face shields or masks, and eye protection and ventilation devices.
xMAP Technology Luminex xMAP Technology microspheres are not monodispersed and tend to aggregate until coated. Multiple pipetting from the original container may affect bead concentrations. Protect the microspheres from light at all times. Follow the resuspension instructions based on the size of container in use: For 1 mL containers: Remove from 2-8°C and allow equilibration to room temperature. Centrifuge for 1-2 minutes at ≥ 8,000 x g.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Repetitive xMAP Microsphere Measurements The reporter, bound to reagents at the surfaces of the xMAP microspheres, provides raw analytical data. Because an xMAP microsphere suspension provides near liquid phase reaction kinetics, each microsphere of a particular signal theoretically binds an equal number of reporter molecules. Equal binding results in a statistically even distribution of reporter on each microsphere in a set.
xMAP Technology Luminex xMAP Technology The volume restrictions that the amount expelled back function places on the assay design can be expounded by the following formula. [“Total well volume” (uL ) – “Sample uptake volume”(uL)] + 160(uL)
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Gating xMAP Technology Determine the gating on the doublet discriminator channel for the assay during development. The numeric values appear on the lefthand side of the histogram. Use the numerical gate position determined in development to set the gate location in the templates. Gating information may change with a new lot of microspheres. Each time you receive a new lot of microspheres, re-evaluate them with the current templates.
xMAP Technology Luminex xMAP Technology In the Create Products section, you include standards and controls for use with your kits. Finally, you define template information that includes sample volumes, how many tests are included in the kit, and analyzer functions. Acquisition Detail Tab The acquisition detail is useful if you are optimizing assays before giving them to end users. You can find the acquisition detail on the Acquisition Detail tab.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Main Controls and Batch Data Click Autosize to expand compressed fields The top half of the screen displays the main control and session detail information. Select Statistics Batch Data Area Acquisition Detail Tab Click the Maximize button to see full list of data Figure 1. Acquisition Detail Screen Displaying Batch Data Use the Acquisition Detail tab to create a New Advanced Batch and dynamically monitor its progress.
xMAP Technology Luminex xMAP Technology Figure 2. New Advanced Batch—General Tab 2. Fill in the Name, Description, and Operator fields. 3. Edit the Sample Size field if desired. The default value is 50 uL. This field accepts values from 20 to 200. 4. Edit the DD Gate field as desired. The default value is 7500 to 15000. The field accepts a range of 0 to 32767. 5. Edit the Timeout field as desired. The default value is 25 seconds. The field accepts a range of 0 to 400 where 0 is no timeout.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Figure 3. New Advanced Batch—Bead Set Tab 8. Edit the Events and Caption field for each bead set. Edit the Default Events field to change the default value. Click Apply to All to apply the default value to all bead sets. Bead Set Tab—Selection Shortcuts Selected Column Select several rows, right-click the Selected column, and then select/ deselect the desired rows.
xMAP Technology Luminex xMAP Technology Bead Set tab. The default value is 1000. The recommended minimum is 100 per bead set. • By selecting Per Bead the system will continue to analyze until each bead set has met the events as determined on the Bead Set tab. • By selecting Total Beads the system will continue to analyze until the total bead event measurement is met. 11. Click the Plate Layout tab. The Options Plate Layout tab dialog appears.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Table 4. Plate Layout Tab—Selection Shortcuts Click column heading to select entire column. Click row heading to select entire row. Click the top left corner of the plate to select entire plate. Each well can have a series of Off-Plate commands that run before the next well is read. You can insert preliminary Off-Plate commands that run before the first well. Figure 5. New Advanced Batch—Popup Command Menu 14.
xMAP Technology Luminex xMAP Technology View Batch Data To open an existing batch previously stored in the data base: 1. Click the View Batch Data button. The Open Batch dialog appears. Note: To redisplay the Histogram, Bar Graph, and Bead Map visuals, raw data must be stored at the time the data is collected. To enable raw data storage, select the Tools menu and select Options. Make sure ‘Enable Raw Data Storage’ is checked on. Start Figure 6. Open Batch 2.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 This does not change the format of the expected data. %CV Count Mean Median* Peak StdDev Trimmed %CV Trimmed Count Trimmed Mean Trimmed Peak Trimmed StdDev *Only use the Median statistic for calculations with the Luminex 100 IS software. The Luminex 100 IS and the Luminex 100 IS Developer Workbench use only Median statistics. Maximize. Click this button to expand the Batch Data to full screen.
xMAP Technology Luminex xMAP Technology Note: The gate that is in effect when collecting data determines which values are used in the result. Applying a gate or changing a gate for existing data does not change your calculated values. The gate positions used while collecting data are the numerical values selected in the template or the New Advanced Batch. Gate boundaries Aggregate Beads Numerical gate position (current values) Figure 7.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology 5. Note the numerical gate position indicated in the parenthesis in the lower left corner of the histogram. Note: You may want to change the X axis of the Dot Plot for troubleshooting purposes. We highly recommend using the default settings in all other instances. 6. Use the numbered gate positions from step 5 in templates and New Advanced Batches for assays with the same buffer composition. 7.
xMAP Technology Luminex xMAP Technology see the Dot Plot, you must use the default axis. To display the bead set information, hover the mouse pointer over the desired region. Hover the mouse pointer over a region to display the bead name Figure 8. Dot Plot Display Example Note: You may want to change the X axis and Y axis of the Dot Plot for troubleshooting purposes. We highly recommend using the default settings in all other instances. To change the Y axis category: 1.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Dot Plot Buttons Use the four buttons at the top of the frame to control the display: Density/Decaying. Use this button to toggle between the default Density Dot Plot and the Decaying Dot Plot. The Decaying Dot Plot feature only plots the 100 most recent events acquired by the Luminex 100 analyzer. This button is only active when viewing the Dot Plot. The Density Dot Plot displays a constant accumulation of events.
xMAP Technology Luminex 100 IS Developer Workbench Software Luminex xMAP Technology After developing an assay, use the Luminex 100 IS Developer Workbench software (Developer Workbench) to create the template file that accompanies the assay kit to the end user. You can access the Developer Workbench software with or without the Luminex 100 IS software running. To start the Developer Workbench: 1. On your desktop, double-click the Developer Workbench icon.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.
xMAP Technology Luminex xMAP Technology Product. A group of standards and controls that you use to create a template. Products are analogous to “reagent kits”. They are used in the template’s definition for the multi-analyte assays that will be performed. Standard. An assay calibrator that is used to generate a standard curve for quantitative testing or to set thresholds for qualitative testing. Use standards to calculate the results of an unknown sample. Template.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology Figure 12. Create New Product Dialog Box 2. Click New to enter a new product into the system. You do not need to press the New button for the first use. 24 PN 89-00002-00-062 Rev.
xMAP Technology Luminex xMAP Technology Select Standard Figure 13. Entering Product Information 3. Select Standard. 4. Enter the Product Number, Product Name, and Manufacturer Name for the product you are creating. 5. Enter the Number of Tests for this product and press Enter key. Each test represents an analyte (the bead ID for each test is selected when you create the template using this product). 6. Enter the Test Name (analyte) and Test Units used for each test.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology The Apply Unit button applies the same unit value entered on the first line to all other tests. For example, if mg/uL is entered for the first Test Unit, then select the Apply Unit button to make all tests mg/uL. 7. Enter the Number of Standards for this product and press Enter key. Note: If the reagent names are similar, click the Apply Suffix button to ensure that each name is unique. 8.
xMAP Technology Luminex xMAP Technology Select Control Figure 14. Creating Control Products 3. Select Control. 4. Enter the Product Number, Product Name, and Manufacturer Name for the product. 5. Enter the Number of Tests for this product and press Enter key. Each test represents an analyte (the bead ID for each test is selected when creating the template using this product). 6. Enter the Test Name (analyte) and Test Units used for each test.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology The Apply Unit button applies the same unit value that you entered on the first line to all other tests. For example, if you enter the mg/uL value for the first Test Unit, then select the Apply Unit button to make all test units mg/µL. 7. Enter the Number of Controls (assay) for this product and press Enter key. Note: If the reagent names are similar, click the Apply Suffix button to ensure that each name is unique. 8.
xMAP Technology Luminex xMAP Technology Select Both Figure 15. Creating Products With Standards and Controls 4. Enter the Product Number, Product Name, and Manufacturer Name for one of the product. 5. Enter the Number of Tests for this product and press Enter key. Each test represents an analyte (the bead ID for each test is selected when you create the template using this product). 6. Enter the Test Name (analyte) and Test Units used for each test.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology The Apply Suffix button applies the number one to the first test name. For each successive test name it applies the test name and number at one increment higher than the previous test name. For example: Suffix = Protein, then Test 1 is Protein 1, Test 2 is Protein 2, and so on. The Apply Unit button applies the same unit value that you entered on the first line to all other tests.
xMAP Technology Create Templates Luminex xMAP Technology There are four template types: • • • • Quantitative Qualitative Data Collection Only Maintenance (no tests) You can create each Quantitative and Qualitative template using one product that is composed of a definable number of standards and controls. Quantitative Assay Using Standards or Controls Note: You must define each Quantitative template using a single product. The product must contain multiple standards.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 3. Select Quantitative for the template type. 4. Enter the Sample Vol (µL) for this template. This is the amount of sample that will be tested. Make the volume in the wells at least 25 µL greater than the sample volume. Note: Templates have a default sample timeout of 25 seconds. The timeout begins when the stream begins.
xMAP Technology Luminex xMAP Technology This list shows all saved products you created. Select the desired product from the list Figure 18. Opening Products Note: If the desired product is not present in the list, select New to create a new product. 9. Highlight the product that you want added to the template. The test and reagent information appears for the highlighted product to help you make your selection. You cannot edit this information here. 10.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Select Bead IDs Figure 19. Selecting Bead IDs 13. In the Min Beads field, enter the number of beads to collect for each test. The Apply Min Beads button applies the same min bead requirement that you entered on the first line to all other tests. 14. Click a Curve Fit field to see a list of formulas. Select one of the curve fit formulas for each test. The available curve fit formulas are listed in.Table 7, “Curve Fit Formulas” .
xMAP Technology Luminex xMAP Technology Select a Curve Fit Formula Figure 20. Selecting Curve Fit Formula If desired, you can use the Apply Curve Fit button to apply the same formula, that you selected on the first line, to all other tests. 15. Click Next to move to the next Template Setup Wizard dialog box: Figure 21. Selecting Template Commands PN 89-00002-00-062 Rev.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology 16. Select commands from the Available Commands list and move them to the Template Commands list using the single arrow pointing towards the template command button. These commands become part of the template and are always performed in the order specified for any batch that uses this template. Each well command is automatically assigned to the next available well.
xMAP Technology Luminex xMAP Technology Table 8: Available Commands (Continued) Command Qualitative Assay Using Standards or Controls Note: You must define each Qualitative template using a single product. The product must contain one standard. The standard is assigned lot value information for each test. A control product is optional. PN 89-00002-00-062 Rev. A Definition Calibrate CAL2 Acquires calibration beads from a well to calibrate the Reporter channel.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Select Qualitative Figure 22. Template Setup Wizard Dialog Box 2. Fill in the general information about your company, the author of the template, the name and version for the template you are creating, and a brief description.
xMAP Technology Luminex xMAP Technology 7. Click Next. The next Template Setup Wizard dialog box appears. Select Apply Standard/ Control Figure 23. Applying Standards or Controls 8. Click Apply Standard/Control to add a product. The Open Reagent Product dialog box appears: PN 89-00002-00-062 Rev.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology This list shows all saved products. Select the desired product from the list Figure 24. Open Reagent Product 9. Highlight the product to add to the template. The test and reagent information appears for the product highlighted to help you make your selection. You can not edit the information here. Note: If the desired product is not present in the list, select New to create a new product. 10.
xMAP Technology Luminex xMAP Technology Figure 25. Selecting Bead IDs 13. In the Min Beads field, enter the number of beads to collect for each test. The Apply Min Beads button applies the same min bead requirement that you entered on the first line to all other tests. 14. Select one of the qualitative formulas shown. Figure 26. Selecting Qualitative Formula PN 89-00002-00-062 Rev.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Table 9: Algorithm Formulas Formula Name Lum Qual Formula Algorithm FI sample -----------------------× Ki FI s tan dard Adv Qual Formula FI sample -----------------------× Ki FI s tan dard Lum Script Formula User defined scripting ability Default Values Default Ranges 0 to .9 .9 to 1.1 1.
xMAP Technology Luminex xMAP Technology Figure 27. Selecting Template Commands 16. Select actions from the Available Commands list and move them to the Template Commands list using the single arrow pointing towards the template command list. These commands become part of the template and will always be performed in the order specified for any batch that uses this template. Each well command is automatically assigned to the next available well.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology To create a template for Data Collection Only: 1. Click the Create Template button on the toolbar. The Template Setup Wizard dialog box appears: Select Data Collection Only Figure 28. Selecting Data Collection Only 2. Fill in the general information about your company, the author of the template, the name and version for the template you are creating, and a brief description.
xMAP Technology Luminex xMAP Technology In the Tests field, enter desired number of tests. Figure 29. Entering the Number of Tests 8. Select the Tests field and enter the number of desired tests and then press Enter key. A blank entry for each test appears in the adjacent area (each test represents an analyte bound to a unique bead set). 9. Enter the Name, Units, and Description fields for each test. The Apply Units button applies the same unit entered on the first line to all other tests. Figure 30.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology 10. Select the Bead ID for each test. Type the bead ID or select it from the drop-down chart (optionally, select the first bead ID then click the Apply Bead ID Increment button to auto fill the remaining entries with sequential bead IDs).: Figure 31. Selecting Bead IDs 11. In the Min Beads field, enter the number of beads to collect for each test.
xMAP Technology Luminex xMAP Technology Figure 32. Selecting Template Commands 13. Select commands from the Available Commands list and move them to the Template Commands list using the single arrow pointing towards the template command button. These commands are then part of the template and are always performed in the order specified for any batch that uses this template. Each well command is automatically assigned to the next available well.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Maintenance Template Create a Maintenance Template for an easy startup procedure, troubleshooting, or calibration. Make sure that any maintenance template you create is consistent with the Luminex 100 analyzer operations and maintenance requirements. See the Luminex 100 Integrated System User Manual Version 2.1 for more information. To create a Maintenance template, follow these steps: 1.
xMAP Technology Luminex xMAP Technology Figure 34. Selecting Template Commands 5. Select commands from the Available Commands list and move them to the Template Commands list using the single arrow pointing towards the template command button. These commands are then part of the template and are always performed in the order specified for any batch that uses this template. Each well command is automatically assigned to the next available well.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Template Maintenance xMAP Technology The system lets you edit and delete existing templates. You can also print directly from this screen if desired. Just select the template and click the Print button. Follow the on screen dialogs. If a batch is associated with a template, then the template ID is shown in light blue. You can only edit these templates if they are given a new version number.
xMAP Technology Luminex xMAP Technology Figure 36. Select Template Information 3. Locate the information that you want to change using the Next and Back buttons at the bottom of all the Template Setup Wizard screens. 4. Change the template information or settings and click Next, then click Save. Click Close to exit. The system saves the changes that you make. Edit a Used Template To edit existing templates that do have batches associated with them.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology Figure 37. Select a Used Template to Edit 2. Select the template to edit (the template ID cell is blue) and click Edit. The Template Setup Wizard dialog box appears showing the selected template. Figure 38. Select Template Information 3. Define a new Template Name and assign a new Version number to the template. 4.
xMAP Technology Luminex xMAP Technology 5. Change the template information or settings and click Save, then click Close to exit. The system saves the changes. Delete an Unused Template To delete templates that do not have batches associated: 1. Click Edit Template. An Open Template dialog box appears. The template ID is white if no batches are associated. If the template has a blue template ID field, you can not edit or delete it.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Delete a Used Template xMAP Technology To delete templates that do have batches associated to them. If the template has batches, the Delete button for that entry is grayed out. In this case, use the Delete Batch button on the left side of the screen. Therefore, first check the batches, then you can delete the template. The template ID cell is blue if batches are associated. Any batch results associated to this batch are deleted from IS.
xMAP Technology Print Template Information Luminex xMAP Technology The system lets you print information and details about a template. There are two ways to print templates. The first method is the easiest and preferred method. The second method allows you to print a template during or after edit or creation. To print template information, follow these steps: 1. Click Edit Template. The Open Template dialog box appears. Figure 42. Select Template To Print 2.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology Figure 43. Selected Template Information 2. Select template and click Edit. Edit the desired template fields. Click Next. The next Template Setup Wizard dialog box appears. The Print button is greyed out and not usable unless you Save during the edit or create process Figure 44. Print a Template 56 PN 89-00002-00-062 Rev.
xMAP Technology Luminex xMAP Technology 3. Click Print at the bottom of the Template Setup Wizard screen. A Template Report dialog appears showing information regarding the open template. Figure 45. Template Report 4. Click the Print icon at the top of the Template Report dialog box. The Print dialog appears. Figure 46. Print Dialog Box PN 89-00002-00-062 Rev.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology 5. Select the desired printer and other options. Click Print. 6. Click the X at the top-right corner of the Template Report dialog box to close the dialog box. Import a Template To import a template: 1. Click the Import Template button on the toolbar. The Import Template dialog box appears listing the available templates with the idt extension. Select a template to import and click Open: Figure 47.
xMAP Technology Luminex xMAP Technology To export a template, follow these steps: 1. Click the Export Template button on the toolbar. The Export Template dialog box appears listing the available templates. Figure 48. Export Template Dialog Box 2. Select the template to export and click Export.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Help About Developer Workbench Information about the Developer Workbench: 1. Click Help on the Developer Workbench Toolbar. 2. From the drop-down menu select About Developer Workbench. Figure 50. Help Menu 3. An About Developer Workbench dialog box appears listing the Developer Workbench name, version, and copyright information. Version 2.1.26 Figure 51. About Developer Workbench Dialog Box 4. Click OK.
xMAP Technology Luminex xMAP Technology Figure 52. Developer Workbench System Information 4. Click on the “X” (upper right corner of the box) to close the System Information box. On-line Help On-line help is available at any time. 1. Click Help on the Developer Workbench Toolbar. 2. From the drop-down menu select Help Contents. Figure 53. Selecting On-line Help 3. From the opening help screen select the desired topic from Contents in the left pane or follow the topics in the right pane.
Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 xMAP Technology Figure 54. Open Help Screen Customer Support 62 Toll free customer support is available to users in the U.S. and Canada by calling 1-877-785-BEAD (-2323) between the hours of 7:00 a.m. and 7:00 p.m. central time, Monday through Friday. Users outside of the U.S. and Canada can call us at +1 512-381-4397 between the hours of 7:00 a.m. and 7:00 p.m. central time, Monday through Friday.
A Compatibility with Luminex 100 Version 1.7 If you are creating assays with the a Luminex 100 Version 1.7 system to run on a Luminex 100 IS system, perform the settings in this appendix. Note: You cannot have Luminex 100 Version 1.7 and Luminex 100 IS loaded on the same system. Record your gate values on the Luminex 100 Version 1.7 system and then enter the information into templates on the Luminex 100 IS system. Certain settings in the Luminex 100 Version 1.
Luminex 100 IS Developer’s Guide to LabMAP Technology Version 2.1 xMAP Technology Figure 55. Options Dialog Box 3. Click New Session on the main screen. From the New Session screen click Settings on the toolbar. On the General tab of the Settings dialog box, set Sample Size to a size between 20 and 200 µL. Set Flow Rate to Fast. Click OK. Figure 56. Settings Dialog Box 64 PN 89-00002-00-062 Rev.
B xMAP Protocols Updates or additions to these protocols are posted on the Luminex website at http://www.luminexcorp.com. At he home page, select select Customer Support.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 sonicating are not recommended for the larger vials as they are not effective in resuspending the microspheres or preventing aggregation. Note: Inserting a sonicator probe into preparations is not recommended. Probe Sonicator Over time, it is normal to observe a presence of beads on the surface of the liquid in the stock vials. You may also notice a container wall residue and suspended macrobead aggregates.
xMAP Technology It is not advisable to construct an assay without defining the concentration of the subject-coupled microsphere preparation. The total surface area (total number of xMAP microspheres) represented in an assay is a critical variable, so the optimization and control of this variable begins with manipulation of known microsphere numbers. xMAP microspheres can be counted with a hemacytometer. Please follow counting and calculation methods outlined in the instructions for your hemacytometer.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 Two-Step Carbodiimide Coupling of Protein to xMAP Carboxylated Microspheres Introduction Use the protocols included here as a general starting point for developing assays. All assays should be optimized for your reagents in your specific application. You will discover the methods that give you the best results by starting with these guidelines and modifying them for your specific needs.
xMAP Technology Preparation 1. Allow all reagents to warm to room temperature. 2. Dilute the protein stock with COUPLING BUFFER to a concentration of 25 - 250 µg/mL and a volume of at least 500 µL (See Technical note 2). 3. Using an analytical balance, weigh approximately 10 mg of Sulfo-NHS into a tube. Repeat for EDC (See Technical note 1). Procedure Microsphere activation 1. Centrifuge the xMAP microsphere stock for 1 minute at ≥ 8,000 × g. 2.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 13. Add 500 µL of COUPLING BUFFER to the activated microspheres and vortex. Centrifuge the activated microspheres for 1 minute at ≥ 8,000 × g. Aspirate the supernatant. 14. Repeat the previous step and go immediately to the protein coupling steps. Coupling, blocking, and storage 1. To initiate the coupling reaction, add 500 µL of the protein preparation (protein stock in COUPLING BUFFER) to the activated microspheres; vortex.
xMAP Technology One-Step Carbodiimide Coupling of Oligonucleotides to xMAP Carboxylated Microspheres Introduction Use the protocols included here as a general starting point for developing assays. All assays should be optimized for your reagents in your specific application. You will discover the methods that give you the best results by starting with these guidelines and modifying them for your specific needs.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 4. Remove the supernatant, being careful not to disturb the pellet. 5. Add 50 µL of 0.1 M MES (pH 4.5). Vortex and sonicate. 6. Add 0.2 nmole of amino-substituted oligonucleotide (that is, 2 µL of a 1:10 dilution of the 1 mM solution). Vortex briefly (See Technical note 3). 7. Immediately before use, prepare a 10 mg/mL solution of EDC powder (See Technical note 4). Vortex until dissolved. 8. Add 2.
xMAP Technology 3. The optimal coupling concentration for a given oligonucleotide is determined by coupling at various concentrations within the recommended range of 1 to 50 µM. 4. Minimize the exposure of EDC to air; secure closures on stock and dry aliquot containers, store desiccated at -20°C. Use aliquots immediately and discard containers after use. Make a fresh 10 mg/mL EDC solution before each addition.
xMAP Technology Luminex 100 IS Developer Guide to xMAP Technology Version 2.1 2. Disperse the pellet with sonication, and vortex the container for 20 seconds. 3. Dispense 1.0 × 105 Avidin-modified microspheres into a 1.5 mL microcentrifuge tube. 4. Centrifuge the microspheres for 1 minute at ≥ 8,000 × g and remove the supernatant. 5. Add 250 µL of the REACTION BUFFER (See Technical note 2). Vortex. 6. Add 250 µL of the Biotin-conjugated molecule solution to the microsphere suspension. Vortex immediately.
Index A About Developer Workbench 1-60 about this guide 1-1 accuracy and precision specifications 1-2 Acquisition Detail screen 1-9 Acquisition Detail Tab, accessing 1-9 Advanced Batch,definition 1-10 Apply Suffix button 1-25 Apply Suffix button,button, Apply Suffix 1-25, 1-27, 1-29 Apply Unit button 1-26 Apply Units button 1-25, 1-27, 1-29 Assay Controls 1-26 assay controls, group 1-26 Assay Standards 1-23 Assay Standards and Controls 1-28 assay standards and controls, group 1-28 Autosize 1-15 autosize 1
xMAP Technology Luminex 100 IS Developer Guide to Technology Version 2.
xMAP Technology template, delete 1-53, 1-54 template, edit 1-50, 1-51 template, export 1-58 template, import 1-58 U user manual, conventions 1-1 V value settings, Luminex 100 Version 1.7 A-63 value settings,Luminex 100 A-63 value settings,Luminex 100 IS A-63 View Batch Data 1-15 vortexing B-65 W warnings biological 1-4 PN 89-00002-00-062 Rev.
Luminex 100 IS Developer Guide to Technology Version 2.
