Applied Biosystems SOLiD™ 4 System SETS Software User Guide ...

Parameter Description
Applied Biosystems SOLiD 4 System SETS Software User Guide
Chapter 2 Prepare Run Settings
Set analysis parameters 2
HQ Beads When visualizing the beads on a satay plot, those that are on the axis and far
from the origin are the beads that have a bright signal in only one channel.
High quality beads are a summary of the “good” and “best” beads that are
reported in the Cycle Scans Report (available in both ICS and SETS).
There are two concept definitions necessary: intensity and angle.
The concept of intensity is defined as the magnitude of the scaled intensity
vector;
The concept of angle is defined as 1 - cos(theta), where theta is the angle (in
four-dimensional scaled intensity space) between the bead intensity vector
and the axis of the called color.
The intensity value can increase due to the total intensity across all channels
independent of the intensity of the primary channel alone.
Considering the angle and the intensity, a “good” bead is a usable bead that
satisfies the following conditions:
intensity >0.05
0.0125 < angle ≤0.05
A “best” bead is a usable bead that satisfies these conditions:
intensity >0.05
angle ≤0.0125
A “best” bead is not a “good” bead because these are exclusive classes.
The set of high quality beads, for this metric, is the merging of the two
classes.
These values are also highly correlated to the quality values (QV) for calls. A
drop in the percentage of beads falling into the HQ category indicates that an
accurate color calling is becoming more difficult.
Bead Retention The number of usable beads in the analysis of a panel for any particular
ligation cycle is nearly always smaller than the total number of beads found in
the corresponding focal map image.
Bead retention is simply the fraction of the total number of beads that are
usable.
Because the stage can drift slightly from one slide scan to the next, beads
near the edges of a panel might not be imaged in all four colors.
The software will call a color only for bead locations that are imaged in all four
color-scans of a ligation cycle (i.e. any usable bead).
Since beads are identified, or found, only during the focal map analysis stage,
a call is made for such a bead location, even if the bead has physically moved
from that position. In such cases, since the bead location is most likely now
vacant, subsequent color calls will be based on data dominated by noise or, if
present, a close neighbor bead. As such beads are still classified as usable,
bead retention is not sensitive to such dislocations.
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