Eppendorf BioSpectrometer fluorescence - Manual (Page 91)

91
Evaluation procedure
Eppendorf BioSpectrometer ® fluorescence
English (EN)
The application of the evaluation procedure can be activated in the parameters Correct A260 or Correct
A280.
A
XXX, corr
= calculated corrected absorbance for a wavelength of 260 nm or 280 nm
A
XXX
= measured absorbance for a wavelength of 260 nm or 280 nm
CF = correction factor for a wavelength of 260 nm or 280 nm (the correction factors for 260 nm and 280 nm
are both dye-specific and are programmed in
General Method Parameter: Dyes in the Functions area).
A
YYY
= measured absorbance at the dye wavelength.
12.5.2 Ratios A260/A280 and A260/A230.
Application: Information on the purity of the measured nucleic acid. Application of the evaluation
procedure can be activated in the
A260/280 or A260/A230 parameters.
"Ratio" refers to the quotients of the measured absorbances at the listed wavelengths.
Literature values for ratio values with pure nucleic acids:
A260/A280
• DNA: 1.8 to 1.9
• RNA: 1.9 to 2.0
(Current Protocols in Molecular Biology, 1994)
A260/A230
For the ratios A260/A230, different information can be found in the literature for pure nucleic acids:
• DNA: 2.3 to 2.5
(The Nucleic Acids, 1955)
• DNA: 1.9
(Current Protocols in Molecular Biology, 1994)
The values are highly dependent on the pH value. Therefore, nucleic acids should not be measured in
water, but in a buffer with a pH of 7 to 7.2 (e.g., TE buffer).
The absorbance values displayed in the results are the directly measured, not the corrected
absorbance values.
YYY XXX corr XXX
A CF A A u
,