Bio-Rad Human Inflammation Assays - Manual (Page 28)

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6. Click Enter Sample Info and enter sample information and the
appropriate dilution factor.
7. Click Run Protocol and confirm that the assay settings are correct.
a. Refer to Table 12 for the recommended RP1 (PMT) setting.
Protocols using alternative PMT settings should be validated by
the end user.
b. Confirm that data acquisition is set to 50 beads per region.
In Advanced Settings, confirm that the bead map is set to
100 region, the sample size is set to 50 µl, and the doublet
discriminator (DD) gates are set to 5,000 (Low) and 25,000 (High).
In Bio-Plex Manager software versions 4.0, 4.1, 4.1.1, and 5.0,
check Override Gates and set the DD gate values as indicated.
Select Start, name and save the .rbx file, and begin data
acquisition. The Run Protocol pop-up screen will appear. Click
Eject/Retract to eject the plate carrier.
Acquire Data
1. Shake the assay plate at 850 ± 50 rpm for 30 sec and visually
inspect the plate to ensure that the assay wells are filled with buffer.
Slowly remove the sealing tape and any plate cover before placing
the plate on the plate carrier.
2. Click Run Protocol and on the pop-up screen select Load Plate
and click OK to start acquiring data.
3. Use the Wash Between Plates command after every plate run
to reduce the possibility of clogging the instrument.
4. If acquiring data from more than one plate, empty the waste bottle and
refill the sheath bottle after each plate (if HTF are not present). Select
Wash Between Plates and follow the instructions. Then repeat the
Prepare Protocol and Acquire Data instructions.
5. When data acquisition is complete, select Shut Down and
follow the instructions.