Agilent Technologies 8453 Operator's Manual page 622

3 Thermal Denaturation Mode
The Use Wavelength at: field allows you to define the wavelength of your
processed data from which the amplitude values are extracted for the
wavelength result. The default wavelength is 260 nm. If you enter a wavelength
at which no measurement is available (for example a non-integer value), the
amplitude values are calculated by linear interpolation.
The Background Correction field allows you to specify any background
correction that is applied to the wavelength result to calculate a function
result. Choose the down arrow and select a background correction procedure
from the drop-down list. The available options are identical to the procedures
in the kinetic mode.
The Temperature group allows you to select the source of your temperature
measurement. Choose the down arrow and select the temperature source from
the drop-down list. If you choose internal sensor the temperature of the
thermostattable cell holder, which is measured by the Peltier temperature
controller, is used for the data evaluation. If you choose external sensor the
temperature of your sample, which is measured by the external temperature
sensor, is used for the data evaluation.
The Set Calculation Range group allows you to limit the calculation of the
results on all traces to a specified temperature range. You specify the lower
and upper limits of the temperature range in the From and To fields.
The Absorbance Ratio group allows you to specify a temperature at which
absorbance data are normalized; the temperature trace is divided by the
absorbance value at the specified temperature. Select Normalize at
temperature to switch on the normalization, and specify the normalization
temperature in the adjacent field.
The Tm Calculation group allows you to select the calculation method for the
melting temperature.
• Choose Average (mean value) to calculate the melting temperature as the
• Choose Derivative to calculate the melting temperature using the first
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temperature at the average value of the absorbance at the start of the
calculation range and that at the end of the calculation range.
derivative of the temperature trace. Enter a Filter length for the derivative
calculation and a Sensitivity for the reporting of Tm values in the adjacent
fields.
Biochemical Analysis Software for Agilent ChemStation