Perkin-Elmer fluorescence detector

Some commercially available detectors have a number of detection modes built into a single unit. Fig. 2.4o is a diagram of the detector used in the Perkin Elmer 3D system, which combines uv absorption, fluorescence and conductivity detection. The uv function is a fixed wavelength (254 nm) detector, and the fluorescence function can monitor emission above 280 nm, based on excitation at 254 nm. The metal inlet and outlet tubes act as the electrodes in the conductance cell. The detection modes can be operated independently or simultaneously, using a multichannel recorder. In the conductivity mode, using NaCl, a linear range of 103 and a noise equivalent concentration of 5 x 10 8 g cm-3 have been obtained. 

Methods. Absorption spectra were recorded using an Hitachi model 150-20 spectrophotometer/data processor system. Uncorrected steady-state fluorescence emission spectra were recorded using a Perkin-Elmer MPF-44A spectrofluorimeter. These spectra were collected and stored using a dedicated microcomputer and then transferred to a VAX 11/780 computer for analysis. Fluorescence spectra were corrected subsequently for the response characteristics of the detector (21). Values of the fluorescence quantum yield, <) , were determined relative to either quinine bisulfate in IN H2S04 )>f =... 

Measurements were performed on a Waters 470 HPLC fluorescence detector equipped with a JASCO cuvette accessory and connected to a Perkin Elmer 561 strip chart recorder. Excitation and emission band-widths were 18 nm. Emission spectra were measured for the three excitation wavelengths mentioned above and emission starting from 10 nm higher than excitation up to 700 nm. Fluorescence at fixed wavelengths was measured four minutes after cuvette insertion and expressed as per-millage of the 275/303 fluorescence of 3.0 pM tyrosine in 50 mM HEPES, pH 7.4. Corrections were made for buffer- and blank collagenase fluorescence, and for signal attenuation. 

Absorption and Fluorescence Instrumentation. Absorption spectra were obtained using a Princeton Applied Research Corp. (PARC) Model 1208 polychromator, a Perkin-Elmer 8 yL absorption flow-cell and a 50 watt deuterium light source. Fluorescence spectra were obtained using a Farrand Mark 1 Spectrofluorometer (previously described (13)) and either a 10 iiL Farrand micro flow-cell, or a Precision Cells, Inc. (Model No. 8830) 20 yL flow-cell. A PARC Model 1254 SIT detector, having a UV scintillator, was mounted on both the absorption polychromator and fluorescence spectrofluorometer. Spectral coverage in the absorption and fluorescence modes was 60 and 115 nm, respectively. All absorption and fluorescence spectra were obtained in one second, i.e., 32 scans of the SIT target. 

Separation of the Priority Pollutants Monitored by the Simple Fluorescence Detector Courtesy of the Perkin Elmer Corporation... 

All HPLC analyses employed a Perkin-Elmer LS-4 fluorescence detector operating at excitation and emission wavelengths of 296 nm and 326 nm respectively. 

The HPLC was a Du Pont 8800 quaternary solvent system with a Hewlett-Packard (HP) 1040A photodiode array detector (5-8). Flexible disks were used for data storage, and postrun data evaluation was performed by the detector s computer (HP 85). Samples were injected with a loop injector 10 p,L was used for analytical scale, and 200 jlL was used for preparative scale. Spectra were run on a spectrophotometer (Perkin-Elmer Lambda 3) for static absorbance and on a spectrofluo-rometer (Perkin-Elmer MPF-66) for fluorescence. Field-ionization mass spectra were obtained at a resolution of 45,000 (corresponding to a 0.01-daltons error for a mass of 450). 

Most FCS setups therefore use single photon avalanehe photodiodes [323,424], usually SPCM-AQR detectors from Perkin Elmer [408]. These deteetors have a quantum efficiency that reaches 80% at 800 nm. However, single photon APDs often have a timing delay and transit time jitter dependent on wavelength and count rate. The changes can be of the order of 1 ns. Recording a fluorescence decay curve under this condition delivers questionable results. 

UV/visible spectra and kinetic data were collected on a Perkin-Elmer 559A spectrophotometer or a Perkin Elmer Lambda 40 spectrophotometer. Product yields were determined on a Perkin-Elmer LC-235 HPLC equipped with a 235C diode-array detector measuring absorbance at A. = 220 nm and a LC 240 fluorescence detector, (ex. X = 210 nm, em X = 330 nm). The HPLC was fitted with a Rainin Microsorb-MV C-18 reverse phase column (4.6 mm ID x 25 cm 5-nm particle size) a 200 pi sample loop. The mobile phase for product separation was 80% MeOH/20% H2O with a flow rate of 0.8 ml/min for ail runs. Reported peak areas are averages of triphcate injections. Solution pH was measured with an Orion combination pH microelectrode, model 8103. a spin-lattice relaxation times, Ti, were measured on a Varian VXR-200 and... 

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