Bandpass detection

From Fig. 4 it can be seen that, for finite bandpass detection, one will obtain different fluorescent intensities per emitting molecule depending on the level pumped. This can produce systematic errors in both the determination of absolute concentrations and the use of excitation scans to obtain ground state rotational temperatures (21) Also, the lack of a thermal distribution imposes restrictions on models of and data analysis in optical saturation techniques. 

There is no sensitivity to individual molecular trajectories or dipole orientations, but one ends up directly with global figures to characterize the emitted fluorescence. Besides, distinguishing between the contributicms of the radiative rate and the collection efficiency remains a challenge, mainly because of the intrinsic difficulty to reliably measure collection efficiency. Lastly, the fluorescence enhancement factors are spectrally averaged within the fluorescence bandpass detection window. However, further investigations can provide some additional knowledge on these last two points, as we will discuss hereafter. 

Quin-2 and PHPA are the only combination we have foimd that allows us to look at Ca and oxidant production simultaneously. For this experiment, Quin-2 is detected at 490 nm through a bandpass filter and PHPA is monitored at 400 nm through the monochromator. Under... 

As we have seen in Section 9.5.3, in the case of resistance thermometry, the signal produced by a low-temperature thermometer is very low (microvolt range). Low-pass filters are not sufficient to narrow the detection bandwidth in order to get a suitable signal to noise ratio (S/N). Bandpass filters are needed. The most commonly used method is the synchronous demodulation, usually simply called lock-in technique, as shown in the block diagram of Fig. 10.7. 

When the emission monochromator of the spectrofluorometer is set at a certain wavelength AF with a bandpass AAF, the reading is proportional to the number of photons emitted in the wavelength range from AF to AF + AAF, or in the corresponding wavenumber range from to 1/AF to 1/(AF + AAF). The number of detected photons satisfies the relationship ... 

Figure 11.15. Schematics of the optical arrangement and temperature probes for the Cr+ fluorescence lifetime-based fiber optic thermometers. F = short-pass optical filter Fa = bandpass or long-pass optical filter LD = laser diode LED = light emitting diode S = the fluorescence material used as sensing element vm = signal to modulate the output intensity of the excitation light source v/= the detected fluorescence response from the sensing element.

Fig. 4. (A) Normalized absorption (solid line) and fluorescence emission (dotted line) spectrum of diglucamide indotricarbocyanine 4 in bovine plasma (concentration 2 pmol/L ) (B) fluorescence image of a tumor-bearing rat (MTLn-3 mammary carcinoma) in anterior view at 0 h and (C) 24 h after administration of 4 (dose 2 pmol/kg body weight), excitation wavelength 740 nm, detection bandpass 750 - 800 nm [45], Copyright 2001 International Society for Optical Engineering (SPIE)...

FIGURE 9.7 Schematic of an in-line interferometer. The anti-Stokes local oscillator field is collinearly overlapped with the pnmp and Stokes beams on a dichroic mirror (DM). All fields are focused by a microscope objective (MO) into the sample (S), and the total signal at the anti-Stokes frequency is detected throngh a spectral bandpass filter (F) at the photodetector. 

For a square-wave-chopped photometer with phase-sensitive detection following a bandpass filter centered on the chopping frequency such that... 

Phosphorescence spectra (uncorrected, front face) were recorded on a Perkin-Elmer LS-5 fluorescence spectrometer using a pulsed excitation source ( 10 ps) and gated detection. The instrument was controlled by a P-E 3600 data station. The samples were typically excited at 313 nm using the instrument s monochromator and an additional interference filter. Excitation and emission bandpasses were 2 nm. Typically the emission spectra were recorded using a 50 ps delay following excitation and a 20 ps gate. The samples were contained in cells made of 3x7 mm2 Suprasil tubing, under a continuous stream of N2, 02 or 02/N2 mixtures of known composition. 

It is also possible to partially alleviate the problem of chemical insensitivity by incorporating narrow bandpass filters into the optical setup.20 Thus, by choosing an appropriate frequency region, it becomes possible to detect the presence of a particular reactant or product species. While this adds some measure of chemical sensitivity to the thermography approach, it is only capable of monitoring one species at a time. Additionally, the success of this approach relies upon the fact that the spectral bands of the desired species do not overlap with any other species and that unexpected reaction products that have spectral contributions in the region of interest are not present. 

Method B. Bisgaier et al. [78] reported a CETP assay using fluorescent cholesteryl 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-3-indacenedodecanoate (BODIPY-CE, Fig. 4) in microemulsions. Microemulsions for donor/acceptor contain triolein, BODIPY-CE/cholesteryl oleate, and l-hexadecanoyl-2-[cw-9-octadecenoyl]-s -glycero-3-phosphocholine. The assay mixtures consist of acceptor microemulsions, donor microemulsions, and a test sample in each well of flat-bottom 96-well plates. After preincubation at 37°C for 10 min, the reaction is initiated by addition of CETP solution. The fluorescent intensity in each well of the plates is periodically (every 10 sec) detected at 37°C with a fluorescent 96-well plate reader equipped with 485- and 538-nm bandpass filters in the excitation and emission paths, respectively. 

In order to integrate the optics in the microchip for fluorescent detection (see Figure 7.7), describe how a bandpass emission filter is fabricated. (2 marks)... 

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