Bio-Rad Experion Protein Analysis Kits - Manual (Page 35)

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5.3 Performing Relative Quantitation
Experion software also automatically performs quantitation of all peaks in a sample by a single-point
calibration to the 260 kD upper marker and presents these data in the Concentration (ng/µl) column of
the Results table.
To view these default relative quantitation results, click on the peak of interest in the electropherogram to
highlight the data for that peak.
To perform relative quantitation against a different, user-defined internal standard:
n
Add the standard to the samples at a single concentration, and
n
Change some of the analysis settings in the software
Selection of the user-defined standard in the software may be done after the run is complete. This
enables easy comparison of the relative quantitation results derived by using both the upper marker and
the user-defined standard.
To perform relative quantitation with a single concentration of a user-defined internal standard:
1. Prepare the protein samples and reagents as outlined in Section 3.5, except add a known
concentration of the internal standard (for example, 500 ng/µl) to each sample. For example, replace
half the volume of sample with the internal standard (combine 2 µl sample, 2 µl internal standard, and
2 µl sample buffer).
Fig . 5 .2 . Internal Std . and Std . Protein
Calibration Curve window.
Do not add the internal standard to the Pro260 ladder.
2. Run the Pro260 analysis.
3. After evaluating the run as described in Section 3.10, select Analysis > Internal Std . and Std
Protein Calibration Curve.
4. In the Internal Std . and Std . Protein Calibration Curve window (Figure 5.2), select Use User
Internal Standard and enter the name, concentration, and molecular weight (MW) of the internal
standard. Click Apply and then Close. The new concentrations appear in the Results table under
the Concentration (ng/µl) column (Figure 5.3).
Experion Pro260 Analysis Kit