Molecular Devices FlexStation II, Operator'S Manual

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3.5 Configuring Instrument Settings in Flex Mode
3.5.1 Wavelengths
You can select the number of wavelengths (wavelength pairs) to be read in the
Wavelengths options dialog box. You can select as many as four excitation and emission
wavelength pairs.
You can select an emission cutoff filter for each pair, or you can use Auto Cutoff.
Selecting more than one wavelength expands the settings box to a maximum of four
wavelengths.
Default settings, or previously specified wavelengths will be displayed. You can choose a
wavelength from the common choices in the pull-down menu or simply type over an
existing setting. You can choose any wavelength within available range.
Scanning monochromator wavelength ranges:
•
Excitation
λ
range = 250–850 nm
•
Emission
λ
range = 360–850 nm
Emission Cutoff Filter Settings
Select an emission cutoff filter that will block as much of the residual excitation light as
possible without unduly reducing the fluorescence signal. The cutoff wavelength options
are 420, 435, 475, 495, 515, 530, 550, 570, 590, 610, 630, 665, or 695 nm. You can
also select None, if you don’t want any cutoff. The cutoff value should be near the
maximum emission wavelength (preferably between the excitation wavelength and the
maximal emission wavelength) but at least 10 nm greater than the excitation wavelength.
Flex mode offers 14 emission cutoff filters. The nominal value of a cutoff filter is the
wavelength at which the filter transmission is 50%. Above that wavelength, transmission
is nearly 0%. In general, you should choose a cutoff filter at a wavelength below the
emission lambda (Em
λ
) maximum of the fluorophore. The goal is to block residual
excitation light and increase the signal-to-background ratio. Sometimes it is appropriate
FlexStation II Operator’s Manual – Rev. D 55