For STarStation 2.3-link to PD 038-301 SS 2.3 operation guide- User Modules on Product page - Section Quick Guides

For STarStation 2.0 Users please refer to the instructions below.

1. Turn on the STarSystem (Note that the XY Platform, Luminex 100 Cytometer and Sheath Delivery Device all have individual power supplies).
-Check for blue lights on the XY platform, Luminex 100 and Sheath Delivery Device (if present).
-Verify that the compressor on the Luminex 100 switches on within 1 minute of the unit being powered up (low rumbling noise).
2. Check that the waste bottle is empty and that the Sheath Bottle or SD System contains sufficient Luminex Sheath Buffer.
3. Start the STarStation Software and login.
4. Launch the STarStation Instrument Controls dialog by clicking the Instrument Controls icon or using the keyboard shortcut (Ctrl + I).

Note that IS version systems begin warming up immediately following power up whereas a warmup must be initiated by the user on 1.7 lx100 systems. This can be confirmed by the Laser Status Indicator on the instrument Status Bar. During warm-up the laser status displays the laser warming icon.
1.7 Version instruments will not warm-up the laser upon powering up and the laser icon will display the laser state as idle.

5. Select the Scripts Tab and select the default startup script. The startup script will warm the detectors, prime the system to remove air bubbles from the sheath buffer inlet tubing and perform an alcohol wash cycle to remove air bubbles from the cuvette.

-Eject the plate holder and place a ACS utility plate on the tray ensuring that the commands outlined in the script are satisfied

R1 - Alcohol (require at least 3ml of 70% ethanol)

Reservoir - Fill with Sheath Buffer.

-Retract the tray.
-Click the Start button on the scripts tab.
-Allow the system to warm up and for the optics to reach operational temperature. This will take 30 minutes (1800 seconds). A timer on the laser status indicator will countdown the remaining warm-up time.

On IS version STarSystems the wash, soak, backflush, prime, drain and Fill and sanitize are able to proceed during laser warm-up, this is NOT possible on a 1.7 version STarSystems.

NOTE: If the system is left idle for 4 hr, the lasers will automatically turn off. A 30 min warm-up period may again be required prior to reading an assay.
NOTE: If the waste is overfilled, the fluidics system may back up and the operation of the system may become less than optimal. The sheath reservoir contains enough fluid for approximately two 96-well plates. If the sheath fluid level falls below the "Sheath" output tubing on the bottle, the assay read cannot be completed.

6. Once the Warm-up, Prime and Wash steps have completed verify that the system pressure displayed on the instrument status bar reads between 6 - 9 psi. Note: the system sheath and air pressures will fall following the completion of an instrument command when the compressor is disengaged. Confirm that the instrument detectors are warm the Laser State icon should display

Probe Height Adjustment- See Technote #40

System Calibration

It is recommended that the system be calibrated at least once a week and whenever the dCal Temperature displayed on the instrument status bar reads +/-3 0c.

The STarSystem calibration status is displayed on the instrument status bar and the status tab of the STarStation instrument controls,

the individual state of the previous reporter and classifier calibrations and the date on which they were performed is also displayed.

Note: the dCal temperature should only used as criteria for deciding whether system calibration is required after the machine has been warmed up and cleansed. Since the ambient temperature of the instrument changes following the system warm-up.

7. If you require a system calibration, ensure that the calibration bead bottles are vortexed prior to use.

Important: Before vortexing, remove the calibration beads from 2-8°C storage and allow the beads to warm to room temperature.

Stage2:Preparing the Assay

Note: this step can be performed whilst the instrument is warming up.

Before you can start collecting sample data you must enter the details for the assay you wish to perform into the STarStation Assay Manager.

1. Ensure you are logged into STarStation under your account name (Assay information once added to the system is only available to the person that entered it).
2. Start the Assay Manager
3. Click the New button.
4. Enter a name for the assay and the number of analytes that the assay detects. Set whether the assay requires the platform heater and the Press the Next button.
5. Enter the name of the analytes detected by the assay and the corresponding bead ID used to detect the analyte.
6. Enter the number of distinct standards and controls that you require for the assay.
7. Click Next.
8. Enter the standard concentration values for each analyte.
Note: Concentration values must be entered from low to high .i.e. standard 1 is the lowest concentration.
9. Enter the concentration values for any controls.
10. Review the assay details and then click the finish button to add the assay to your login account.

Stage 3:Preparing the Plate/Worklist

Note: this step can be performed whilst the instrument is warming up.

1. Ensure that you are logged-in to STarStation under your account.
2. Select the Worklist Tab.
3. From the Multiplex Assay drop-down list select the assay you wish to prepare a plate layout/worklist for.
4. Add standards, controls and unknowns to the plate.
5. Select the direction of sample data collection.
6. Edit any dilution factors and sample id's as required.
7. Give the worklist a name and save it.
8. Select the Close option from the File menu.

Stage4: Reading the Plate

NOTE: Worklists must be created prior to reading a sample. The standard concentrations cannot be added / modified after the plate has been read. Therefore it is imperative that you ensure when creating and selecting Worklists that you assign time to check that the plate layout and assay described in the Worklist accurately reflect the physical setup of the microtitre plate containing your assay.

1. Check that the filter plate is flat. While pressing on one end of the plate, observe the distance that the opposite end of the plate is raised off a flat surface. If the distance is greater than 1 mm, transfer all contents to a flat-bottom 96-well plate or another pre-wetted filter plate and proceed with reading the assay (having ensured optimal probe height).
2. Visually inspect the plate and ensure that corresponding assay wells are filled with buffer prior to placing the plate on the XY platform.
3. Shake the assay plate at 1,100 rpm for 30 sec immediately before starting the run. Failure to do so will result in an increased read time due to settling of the beads. Remove the sealing tape and any plate cover before placing the plate on the XY platform.
4. Select the Acquisition Tab in the STarStation software.
5. From the file menu select the Open option or click the File Open Icon on the file toolbar.
6. Select the Worklist which corresponds with your sample plate from the Worklists directory.

7. After STarStation opens the template worklist, you should verify that the correct worklist has been selected via inspecting that the number of analytes the worklist has been setup to detect corresponds with either the number of analyte tabs displayed in the data grid or the number of bead regions displayed on the classifier plot. If there is a discrepancy in the worklist setup, return to the worklist tab to review or edit the worklist.
8. Press the start acquisition button.
9. The first page of the acquisition wizard is displayed,
- Enable events per bead set (100).
- Enter a sample volume.
- Set the stop time equal to the sample volume (assuming that the sample injection rate is set at 1 microliter/s).

10. Press the Next Button and set the Platform Heater temperature if necessary.
11. Press the Next button and Eject the plate holder and load the assay microtiter plate.
12. Proceed to the final page of the acquisition wizard, Press the Start Data Acquisition Button

13. Once data collection begins use the DD plot and Classifier plot to verify that adequate bead events are detected and that the distinct color coded beads are classified appropriately. When satisfied that sufficient beads are being counted we recommend leaving the system to collect data (modifications to gating or classification can be performed via the list mode data during analysis).

NOTE: It is possible to skip specific wells or analyze a well (sample) a second time using the pause acquisition function of the STarStation software.

Note: If reading more than 1 plate, empty the waste and refill the sheath containers after each plate is run. We also recommend performing an alcohol flush followed by two washes with sheath fluid between each plate and warming the laser.

Stage 5: Preparing STarSystem for Shutdown

1. When the assay reading is complete, select the Scripts tab from the STarStation instrument controls and select the default shutdown script.

2. The shutdown script will decontaminate the system and wash the salt-containing sheath buffer from the sample needle and sample tubing reducing the potential for salt precipitation.
3. Eject the plate holder and place a ACS utility plate on the tray ensuring that the commands outlined in the script are satisfied

R2 - 3ml of 10% household bleach or hypochlorite solution.

R3 - 2ml Distilled Water

4. Click the Start button to launch the script.
5. When the Shutdown script completes the STarSystem can be powered off.

Note: It is recommended that you wash with Distilled water (Soak) at least three times before shutting down the instruments. This step is crucial in preventing the precipitation of salts from the sheath buffer in both the sample tubing and sample probe.