Detector Control
Recording Fluorescence Spectra (FLD-3x00(RS)

With the Dionex FLD-3x00(RS) detectors, you can also record several types of spectra. This allows you, for example, to record spectra for determining the optimum emission and excitation wavelengths. Spectrum scans can be triggered manually by executing a command or in a sample Program.

For details about the available scan modes, refer to the FLD-3x00(RS) Operating Instructions.

 Tips:

During the scan process, no other commands can be sent to the fluorescence detector.

The duration of the scan process depends on the range to be scanned and the selected scan speed.

Before each scan, select detector sensitivity (ScanSensitivity), filter wheel setting (ScanFilterWheel), and scan speed (ScanSpeed).

All scan commands are also available on the FLD control panel (click More Options).

Generally, the scans indicate the best wavelength (pairs) for a specific detector. In order to run a scan for best comparability with spectra in the literature, use the ScanSignalType property and set it to ExCorrected.

Artefacts may occur when scanning near steep flanks, for example, when scanning the emission spectrum of water near the excitation of 350 nm. In this case, select a greater distance between the wavelength scan range and the edge. For the Raman scan, for example, select a start wavelength of 385 nm.

To record a baseline spectrum

It is not mandatory to perform a baseline scan prior to every excitation/emission/synchro scan. However, you must record a new baseline spectrum before the scan whenever you change the scan settings. The baseline spectrum must be recorded before the scan, using the same scan parameters There are two options:

• Record the baseline spectra outside of peaks, but as close as possible to the peak to be scanned to maintain the same conditions.

• In case of gradients, we recommend to run a blank run and perform the baseline scan at the same time the spectra scan is performed. This is only possible if only a single spectra scan (for each scan mode) is performed per sample.

1. Open the Commands dialog box for the detector. Select the scan parameters for your scan mode.

2. Execute the BaseLineScanEmission, BaseLineScanExcitation, or BaseLineScanSynchronous command, depending on your scan mode.

The baseline spectrum is stored and automatically subtracted from the recorded spectra. The result is a difference spectrum. If the parameters for a scan change, the baseline spectrum is automatically deleted and a new baseline spectrum must be recorded. You can delete a baseline spectrum any time by using the ClearBaseLine command.

To record an excitation spectrum

1. Open the Commands dialog box for the detector. Under ScanStartExWavelength, enter the first wavelength of the range to be scanned and the last wavelength under ScanEndExWavelength. Select an emission wavelength (ScanStartEmWavelength).

2. Stop the pump flow (Flow=0) or set it to a very low flow rate (may not be required, depending on application).

3. Execute the ScanExcitation command.

To record an emission spectrum

1. Open the Commands dialog box for the detector. Under ScanStartEmWavelength, enter the first wavelength of the range to be scanned, and the last wavelength under ScanEndEmWavelength. Select an excitation wavelength (ScanStartExWavelength).

2. Stop the pump flow (Flow=0) or set it to a very low flow rate (may not be required, depending on application).

3. Execute the ScanEmission command.

To record a synchronous excitation and emission spectrum (Synchro Scan)

1. Open the Commands dialog box for the detector. Under ScanStartExWavelength, enter the first wavelength of the range to be scanned, and the last wavelength under ScanEndExWavelength. Then enter the offset between the scanned wavelengths under ScanSyncOffset.

2. Stop the pump flow (Flow=0) or set it to a very low flow rate (may not be required, depending on application).

3. Execute the ScanSynchronous command.

Chromeleon automatically saves the spectra to a Spectra Library named FLD3000.LIB. The spectra library is saved in the timebase directory on the local data source. If a scan is performed during sample processing, the library is saved in the associated sequence.

To open a recorded spectrum, double-click the spectra library. The recorded spectra can be viewed in Chromeleon.

For a list of all fluorescence detector commands, refer to  Dionex UltiMate 3000 Series Fluorescence Detectors.