Bio-Rad Bio-Plex System Validation and Calibration Tools - Manual (Page 8)

Dynamic Range of Reporter Channel
Definition
The dynamic range is the calculated number of decades covered by the log
amplifier from the slope and the histogram scale. The available range of channels
on the Bio-Plex array reader is 4.5 log amp decades or 32,767 relative linear
channels. The acceptable dynamic range of fluorescence measured by the Bio-Plex
array reader using the Bio-Plex reporter beads is 4.15–4.28.
Impact on Assay Performance
The dynamic range of the Bio-Plex array reader is 4.5 log amp decades or 32,767
relative linear channels. It is desirable for the range of the instrument to be greater
than the range of an assay. If the dynamic range of the instrument is less than that
of an assay, the range of quantitatable analyte may be limited. This parameter will
guide the user in defining the instrument versus the assay dynamic range
limitations.
Linearity of Reporter Channel
Definition
The reporter validation bead set is utilized to construct a plot where the reporter
channel median fluorescence intensity values are plotted against the corresponding
assigned MESF values. Instrument linearity is expressed as the coefficient of
determination or R-squared (R
2 ) value. The R 2 value must be >0.995.
Impact on Assay Performance
The linearity of the instrument response may directly affect a typical standard or
calibration curve in a Bio-Plex assay, thereby impacting the unknown values
extrapolated from that curve. If the R
2 value is not within acceptable limits, it may
be necessary to realign the optics or check the response of the reporter
photomultiplier tube.
Accuracy of Reporter Channel Response
Definition
The accuracy of the reporter channel response is a more stringent measurement of
the linearity than the R
2 value. Simply stated, the accuracy of the reporter channel
response is the percent difference that the regression line is away from the actual
MESF value data points. The desired accuracy value is >90%.
Impact on Assay Performance
Since accuracy is also a measurement of the linearity of the instrument response,
the same principles that apply to linearity also apply to accuracy of the reporter
channel response. Accuracy values <90% could impact assay performance. The
accuracy data is evaluated in combination with optics alignment to determine if the
Bio-Plex array reader will perform according to specifications. It is possible for the
accuracy value to fall out of specification before the linearity parameter. This is
expected due to the fact that the accuracy parameter is a more sensitive
measurement of linearity than the R
2 value. These data are correlated with optics
alignment data as well as assay performance to determine when the array reader
will not perform according to specifications.
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