Bio-Rad Bio-Plex Software® Upgrades and Conversions - Manual (Page 119)

Plate Loading Guidelines
107
Doublet Discriminator Gates
During a reading, a doublet discriminator (DD) channel measures the amount
of light scatter from particles that flow past the red laser. The light scatter is
directly proportional to particle size, and an internal DD “gate” is designed to
identify particles smaller or larger than a single microsphere, including
microspheres that are clumped or aggregated.
In the Advanced Run Settings dialog, the default DD Gates range for the
100-region map is dependent on the type of bead used. For the 5.5 micron,
non-magnetic, polystyrene beads the gates are 4335–10000, whereas the
gates for the 6.5 micron, magnetic beads are 5000–25000. Some Bio-Rad
assays are built on a larger 8.1 micron, magnetic bead which uses DD gates of
5000–32000.
If you are using different-sized beads or want to change the default settings,
you can change the size of the gate by selecting the Override Gates checkbox
and entering new values in the Low and High fields.
You can also change the range by dragging the red lines on the Run Protocol
window histogram (see page 149).
NOTES:
• The values in the Low and High fields are channel numbers
corresponding to the amount of light scatter from a particle. If you
want to manually change the range, you need to know the
approximate channel numbers corresponding to the size of the
beads you are using
• You can change the DD gate range in the Protocol before a reading
or in the Results file after a reading (see page 113). If you change
the DD gate range in the Results file, a record of the range during
the reading is preserved
• You can set the gates on only the DD channel, not the RP1, CL1, or
CL2 channel
• For Bio-Plex assays or Luminex-type assays from other
manufacturers, the default DD gate settings should work well.
However, if you are developing your own assay, the position of the
DD peak may shift and thus the DD gates may need to be shifted
accordingly. Note that you can adjust the DD gate range after a
reading in the Results file and reanalyze the data
Plate Loading Guidelines
Before starting the run, note the following plate handling guidelines and
warnings:
BioPlex_6.book Page 107 Friday, September 23, 2011 11:55 AM