Bio-Rad Experion Protein Analysis Kits - Manual (Page 53)

How Experion Software Analyzes Proteins
Data Presentation
As the photodiode detects fluorescent signals from the dye-LDS micelle-protein complexes, Experion
software converts the signal into an electropherogram (a plot of fluorescence vs. time). Experion Pro260
electropherograms generally have the following features (Figure A.1):
n
Sample peak(s) — signal(s) generated by the sample protein(s)
n
System peaks — cluster of signals generated by small molecules that interact with LDS
micelles. The system peaks are not considered in the concentration determination
n
Upper and lower markers — signals generated by the internal upper (260 kD) and lower
(1.2 kD) markers, which are included in the sample buffer to normalize the separation of
proteins across all wells in a chip. The upper marker is also used as an internal standard for
relative quantitation (see below). For each lane, comparison of the area of this peak with the
area of every detected peak allows estimation of relative concentration
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Fig . A .1 . Electropherogram generated by Experion Pro260 analysis of immunoglobulin (IgG) under nonreducing conditions .
The relative positions of the lower marker, system peaks, sample peak (IGG), and upper marker are shown.
Once separation occurs, the data are converted into a densitometric gel-like image, or virtual gel
(Figure A.2). Each lane in the virtual gel corresponds to a different sample. The sample and system
peaks and upper and lower markers seen in the electropherogram also appear in the virtual gel.
Upper marker
Lower marker
System peaks
IgG
Experion Pro260 Analysis Kit