Hitachi F 2500 Manual
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A - 23 (2) Operating Procedures (a) Set PMT Voltage to 400 V and Data Mode to “Fluorescence”. Perform “Zero Adjust” to calibrate the zero point. (b) Set the triangular cell filled with Rhodamine B into the cell holder in the sample compartment. (See Fig. F-2). The triangular cell must be set in the direction shown in Fig. F-3. Fig. F-2 Setting of Rhodamine B Fig. F-3 Setting Direction of Triangular Cell (Top view) Excitation light Emission Triangular cell Metal part C915758 Triangular cell with metal fitting 650 to 1640. (filled with Rhodamine B)
A - 24 (c) Select “EX” of “Instrumental Response: Utility”. EM wavelength is set at 640 nm and the instrument starts EX wavelength scan (from 198.6 nm to 601.4 nm). At this stage, the instrument is automatically set in the following conditions. Slit (EX/EM) ........... 5.0/20.0 nm Scan speed ............ 60 nm/min Scan mode............. Fluorescence PMT Voltage .......... 400 V Response............... Auto (b) Upon completing the measurement, all of measuring conditions are set to those set before. Now, instrumental response on the excitation side has been measured and stored. Take out the triangular cell filled with Rhodamine B.
A - 25 F.2 Measurement of Instrumental Response on Emission Side This is the function to obtain the instrumental response on the emission side such as wavelength characteristics of the emission monochromator and detector (photomultiplier). The instrumental response on the emission side is determined by measuring a combination of instrumental response on both the excitation and emission sides by synchronous wavelength scan, and dividing it by the instrumental response on the excitation side preliminarily measured. An emission spectrum is correctable within a range of 200 to 600 nm. (1) Operating Procedures (a) Set PMT Voltage to 400 V and Data Mode to “Fluorescence”. Perform “Zero Adjust” to calibrate the zero point. (b) Set the diffuser into the cell holder in the sample compartment. (See Fig. F-4). Fig. F-4 Setting of Diffuser NOTE: Measurement of instrumental response on the excitation side must be completed before executing a measurement of instrumental response on the emission side. C915759 Diffuser 650-1576
A - 26 (c) Select “EM (200 to 600 nm)” of “Instrumental Response: Utility”. EM wavelength is set at 198.6 nm and the instrument starts synchronous wavelength scan (from 198.6 nm to 601.4 nm). At this stage, the instrument is automatically set in the following conditions. Slit (EX/EM) ......... 5.0/20.0 nm Scan speed .......... 60 nm/min Scan mode .......... Fluorescence PMT Voltage ........ 400 V Response ............ Auto (d) Upon completing the measurement, all of measuring conditions are set to those set before. Now, instrumental response on the emission side has been measured and stored. Take out the diffuser.
A - 27 F.3 Measurement of Instrumental Response on the Long Wavelength Range of Emission Side (only with Model F-4500) When a corrected spectrum in the long wavelength area of 600 nm is required on the emission side, the beam of a lamp for which the wavelength characteristics are known is allowed to be irradiated to the emission optics, and the instrumental response in the long wavelength range on the emission side is read for the correction. As a light source, a sub standard light source (P/N 250-0124 or 250-0125) is used. The correction range by it is 500 to 900 nm and, when combined with an instrumental response in the short wavelength range, a spectrum correction is available over the entire wavelength range (200 to 900 nm) on the emission side. (1) Mounting and Positioning Sub Standard Light Source Refer to the instruction manual for the installation of the sub standard light source (optional accessory P/N 250-0124 or 250-0125). (2) Operating Procedures (a) Close the cover of the sample compartment. (b) Set PMT Voltage to 400 V. Perform “Zero Adjust” to calibrate the zero point. (c) Turn ON the power switch of the sub standard light source and wait for approx. 5 minutes until the lamp stabilizes. (d) Select “EM (500 to 900 nm)” of “Instrumental Response: Utility”. The instrument starts EM wavelength scan (from 498.6 nm to 901.4 nm). At this stage, the instrument is automatically set in the following conditions. Slit (EM) ............... 5.0 nm Scan speed .......... 60 nm/min Scan Mode .......... Luminescence PMT Voltage ........ 400 V Response ............ Auto
A - 28 (e) Upon completing the measurement, all of measuring conditions are set to those set before. Now, instrumental response on the emission side has been measured and stored. Remove the sub standard light source.
INDEX - 1 INDEX A ASCII text file ............................................................................................................. 1-3, 6-25 Activity ....................................................................................................................... 3-16, A-6 Analog output gain ................................................................................................... 1-41, 8-10 Auto pre-scan ................................................................................................................... 1-40 Autoscale Y Axis ............................................................................................................... 7-10 B Bird’s-eye view ........................................................................................................... 1-2, 5-11 Blank measurement ............................................................................................ 4-21, 8-1, 8-9 C CAT (Computing for Averaging Transient) ....................................................................... 2-11 Calibration curve data .............................................................................................. 4-21, 4-22 Calibration curve graph ..................................................................................................... 4-22 Calibration type ............................................................................................................ 4-7, A-1 Concentration unit ............................................................................................................. 4-10 Contour ...................................................................................................................... 5-11, 5-9 Corrected spectra ........................................................................................1-26, 2-9, 5-8, A-22 D Data menu .......................................................................................................... 2-5, 3-4, 4-12 Delay ............................................................................................................ 2-7, 3-6, 4-13, 5-6 Derivative ................................................................................................................... 4-6, 6-12 Description of fluorometry................................................................................................... A-12 Details of quantitation ......................................................................................................... A-1 Details of rate analysis function .......................................................................................... A-5 Determination coefficient ................................................................................... 4-22, A-6, A-7 Digit after decimal point .................................................................................................... 4-11 Display tab ........................................................................................................................ 1-30 E Error message .................................................................................................................. 1-27 Event Log ............................................................................................................................ 7-7 Ex power monitor ..............................................................................................................1-41
INDEX - 2 F Factor input ......................................................................................................................... 4-7 Fonts tab ........................................................................................................................... 1-32 G General tab .................................................................................................... 2-3, 3-2, 4-2, 5-3 Gradient ..................................................................................................................... 3-16, A-6 H Help menu .......................................................................................................................... 7-8 I Info tab .............................................................................................................................. 6-26 Installation ........................................................................................................................... 1-5 Instrument tab .............................................................................................. 2-4, 3-4, 4-11, 5-4 Instrument toolbar .................................................................................................... 1-20, 1-22 Integration method ....................................................................................................... 6-8, A-9 Integration time ................................................................................................................. 4-13 Interrupt measurement ................................................................................................ 8-1, 8-9 J JCAMP-DX file ........................................................................................................... 1-3, 6-25 K K factor ............................................................................................................................. 3-10 L Lamp Off ........................................................................................................................... 1-41 Line points tab .................................................................................................................... 6-6 Luminescence ..................................................................................................... 2-5, 3-4, 4-12 M Measurement mode ....................................................................................... 4-3, 5-3, 2-3, 3-3 Measurement toolbar ........................................................................................................ 1-20 Monitor tab ................................................................................................. 2-10, 3-8, 4-15, 5-8
INDEX - 3 N Number of wavelengths ...................................................................................................... 4-10 P Peak area ........................................................................................................................... 4-5 Peak detection ......................................................................................... 1-3, 2-12, 3-10, 5-10 Peak height ......................................................................................................................... 4-6 Peak table .................................................................. 1-16, 2-18, 2-14, 3-12, 3-16, 5-11, 5-15 Photometry ......................................................................................................... 1-1, 4-1, 7-18 Pre-scan ........................................................................................................................... 2-16 Print Preview .......................................................................................... 2-19, 3-17, 4-24, 5-17 Processing tab ................................................................................................ 2-11, 3-10, 5-10 Processing toolbar ............................................................................................................1-21 Q Quantitation tab .................................................................................................................. 4-4 Quantitation type ................................................................................................................. 4-4 R Ratio ..................................................................................................................................... 4-7 Remeasure ................................................................................................................ 4-25, 8-4 Replicates ........................................................................................................... 2-9, 3-8, 4-13 Report tab .............................................................................................. 2-13, 3-11, 4-16, 5-10 Response .............................................................................................................. 2-8, 3-7, 5-7 Result of rate calculation .................................................................................................. 3-16 S Sample data ...................................................................................................................... 4-21 Sample sipper ..................................................................................................................... 8-1 Sample table ............................................................................................ 2-15, 3-13, 4-18, 7-9 Saved calibration curve .................................................................................................... 4-27 Scale Y Axis Down ........................................................................................................... 7-10 Scale Y Axis Up ................................................................................................................ 7-10 Scale tab ...................................................................................................................... 6-4, 6-5 Scan speed .................................................................................................................. 2-7, 5-6 Scan tab .............................................................................................................................. 7-3 Sens. check ...................................................................................................................... 1-41 Shutter closing .................................................................................................................. 1-40 Shutter control ............................................................................................. 1-26, 2-9, 3-8, 5-8 Shutter opening ................................................................................................................ 1-40 Smoothing .................................................................................................................. 1-3, 6-11
INDEX - 4 Spectral calculation ........................................................................................................... 6-16 Spectrophotometer menu ................................................................................................. 1-40 Spectrum toolbar ..................................................................................................... 1-20, 1-22 Stack ................................................................................................................................. 7-11 Standard ........................................................................................................................... 4-21 Standard deviation ............................................................................................ 1-2, 4-13, 4-23 Standard toolbar ...................................................................................................... 1-19, 1-21 Standards tab ................................................................................................................... 4-14 Statistical calculation ................................................................................................. 1-2, 4-23 T Threshold .......................................................................................... 2-12, 3-10, 4-5, 5-10, 6-8 Tile .................................................................................................................................... 7-11 Time scan ........................................................................................................... 1-1, 4-1, 1-17 Toolbar .............................................................................................................................. 1-19 U User Notes .......................................................................................................................... 7-7 V View menu ........................................................................................................................ 7-10 W Wavelength drive system .................................................................................................. 1-11 Wavelength scan ................................................................................................ 1-16, 2-1, 6-1 Wavenumber ...................................................................................................................... 6-5 X Xenon lamp ....................................................................................................................... 1-11 Z Zero adjust ........................................................................................................................ 1-40