Eppendorf BioPhotometer plus - Manual (Page 22)

5 Operation
22
BioPhotometer plus — Operating manual
5.7 OD 600
With method "OD 600" you can measure the bacteria density via a turbidity measurement at
approx. 600 nm. Because this is a stray-light measurement the result depends on the geometry
of the light path which can differ for the photometers by different manufacturers.
The exact measurement wavelength for the BioPhotometer plus is 595 nm. Measurements with
suspensions of E. coli bacteria (absorbance range: approx. 0.1 to 0.3 E) showed at 600 nm about
1 to 2 % higher absorbance values than at 595 nm. You can program a corresponding correction
factor in the parameters.
Results display
5.8 Dye-labeled biomolecules ("dye methods")
The preprogrammed methods for the keys dye 550 and dye 650 contain measuring procedures
for dye-labeled biomolecules. During these measuring procedures both the biomolecule (nucleic
acid or protein) and the dye are measured at different wavelenghts and their concentrations are
determined. In addition the frequency of incorporation (concentration ratio between dye and
biomolecule) is calculated.
5.8.1 Method group "dye 550"
The dye is measured at 550 nm, the nucleic acid at 260 nm and the proteins at 280 nm (see
Methods on page 16). A selection of 4 dyes is available in the method parameters:
• CY 3
• ALEXA 546
• ALEXA 555
• DYE 550
The first 3 dyes are used more frequently in the laboratory, the last dye (DYE 550) is a
spaceholder for other dyes.
For every dye you have to program the following corresponding data in the parameters. For the
Cy and Alexa dyes values have already been preprogrammed but can be modified:
• Absorbance coefficient in the unit cm -1 ·M -1 . The device uses this to calculate the factor for
converting the absorbance into concentration related to a cuvette optical path length of 10
mm and displays this on the start-up screen after opening the method.
• Optional: factors for the correction calculation "CORR: A
550
" (see below for explanation).
The parameters for the nucleic acid or protein component mainly match the parameters of the
method group DNA and RNA and for the method Protein 280 nm.
5.8.2 Method group "dye 650"
The dye is measured at 650 nm. A selection of 3 dyes is available:
• CY 5
• ALEXA 647
• DYE 650
The additional explanations found under method group "dye 550" apply accordingly.
Top: method name and sample number
Center: result.
Bottom right: measured absorbance.
EN
Oper
at
ing man
u
al