Eppendorf BioSpectrometer fluorescence - Manual (Page 90)

Evaluation procedure
Eppendorf BioSpectrometer
® fluorescence
English (EN)
90
12.4 Dilution
In the measure samples method step. entered dilutions are considered in the result calculation:
C
Dil, corr
= result converted using the dilution factor
V
S
= volume of the sample in the measuring solution
V
Dil
= volume of the diluent in the measuring solution
12.5 Special evaluation procedures for nucleic acids and protein UV
This section covers the evaluation of nucleic acids or proteins in the Nucleic acids and Proteins direct UV
method groups, as well as the corresponding biomolecular components in the Dye labels method group.
12.5.1 Correction A
260
and correction A
280
Application: correction of the influence of dye absorbance on the nucleic acid or protein absorbance at 260
and 280 nm for the methods of the
Dye labels group.
• Use the "linear regression" procedure for calibration lines.
• With curvilinear gradients, test which evaluation procedure (quadratic regression, cubic
regression, spline interpolation) produces the function that is most suitable to the standard
evaluation. Spline interpolation connects the measuring points by cubic polynomials,
whereas the regression methods position a quadratic or cubic function between the
measuring points in such a way, that the smallest possible deviation from the function
results for the measuring points.
• Aside from the calculated regression equation, the regression method also displays the
coefficient of determination as a measure for the scattering of the measuring points around
the calculated function. At a value of < 0.8 for the coefficient of determination the result is
issued with a warning.
• If the first standard hat a concentration of "0", select the setting in which the regression
line (regression curve) goes through the zero point.
• If none of the procedures recommended for curvilinear gradients produce satisfactory
results, select the "linear interpolation" procedure.