Sample Fluorescence

Other methods of reducing the fluorescence background from the Raman spectrum focus on the sample itself. The easiest method, and perhaps the one requiring the greatest degree of hand waving, is the so-called drench-quench technique which involves a prolonged exposure (minutes to hours) of the sample to the full power of the laser beam in order to bleach out the fluorescine centers oresent in the sample matrix. The most 

---

Figure 2. The binding and dissociation of FLPEP and receptor on intact neutrophils at 37 C The data are plotted as the specific binding of FLPEP (pmoles/10 cells) on a log plot versus time. Experimental details 10 cells/mL were exposed at time 0 to 1 nAf FLPEP. At 15, 30, 60, or 120 s, antibody to fluorescein is added to each sample. Fluorescence is monitored continuously during the additions. The data ate derived from a point-by-point comparison of the fluorescence measured under conditions of receptor binding and receptor blockade. Data are representative of observations in more than 10 separate experiments. (Reproduced with permission from reference 22. Copyright 19S7 Journal of Biological Chemistry.)...

Visible lasers are typically used for sample excitation, although near-IR lasers can be used when visible excitation sources cause sample fluorescence, obscuring the Raman scatter. 

Purify the derivative by gel filtration using a PBS buffer or another suitable buffer for the particular protein being modified. The use of a desalting resin with low exclusion limits work well. To obtain complete separation, the column size should be 15-20 times the size of the applied sample. Fluorescent molecules often nonspecifically stick to the gel filtration support, so reuse of the column is not recommended. 

FTRS was not originally used on calcified tissue because of problems with sample fluorescence and low signal-to-noise ratio. However, the introduction of near infrared lasers and improvements in the technique have allowed these... 

The commercially available laser source is a mode-locked argon-ion laser synchronously pumping a cavity-dumped dye laser. This laser system produces tunable light pulses, each pulse with a time duration of about 10 picoseconds, and with pulse repetition rates up to 80 million laser pulses/second. The laser pulses are used to excite the sample under study and the resulting sample fluorescence is spectrally dispersed through a monochromator and detected by a fast photomultiplier tube (or in some cases a streak camera (h.)) ... 

Practically, the phase delay and the modulation ratio mR of the light emitted by the scattering solution (solution of glycogen or suspension of colloidal silica) are measured with respect to the signal detected by the reference photomultiplier. Then, after rotation of the turret, the phase delay r/ F and the modulation ratio mF for the sample fluorescence are measured with respect to the signal detected by the reference photomultiplier. The absolute phase shift and modulation ratio of the sample are then — [Pg.179]

Schmid et al. used the same principle to develop sensors to be incorporated into FI systems for the determination of ascorbic acid in fruit juices [38] and that of lactic acid in dairy products [39]. The membrane used in both applications consisted of decacyclene dissolved in silicone rubber that was treated similarly as the membrane in glucose sensors (Fig. 3.4.B). The oxygen optrode was coated with a sheet of carbon black as optical insulation in order to protect it from ambient light or intrinsic sample fluorescence. Ascorbic acid oxidase or lactic acid oxidase was immobilized by adsorbing it onto carbon black and cross-linking it with glutaraldehyde. The FI system automatically buffered and diluted the food samples, thereby protecting the biosensor from a low pH and interferents. 

For the in situ Raman studies the Mo03/Si02 samples were pressed into wafers at 21 MPa. The pressed wafers were placed in a rotatable sample holder that was enclosed in a quartz cell equipped with an oven. The samples were oxidized at 773 K for 1 hour in O2 to minimize sample fluorescence, then were cooled to 343 K in flowing helium. The adsorbates were introduced through an injection port of the cell at 343 K with flowing helium, and the spectra were recorded at the same tenq>erature. 

Atomic absorption, but possible loss of volatile elements during the treatment of the sample. Fluorescence X would be, essentially, analysis of the surface only. 

Dry Oil Oxidation Fluorescence may be measured within or over an oil or within or over an emulsion of lipid in water providing the pH is below 5.5. When plate fluorescence is to be measured, solid sample fluorescence spectrophotometry is necessary (23). A Hitachi Solid Sample Holder Attachment for Model MPF-2A Hitachi-Perkin Elmer Fluorescence Spectrophotometer (Figure 4) was... 

NEXAFS experiments on NOM can be conducted in several modes that differ in the type of detected particle and objectives of the experiment transmission (X rays transmitted through the sample), fluorescence (fluorescent X rays due to absorption of the X-ray beam), or electron yield (photo-emitted electron) (Sparks, 2003). Alternatively, the techniques can be divided into full-field applications such as transmission X-ray microscopy (TXM) and X-ray photoemission electron microscopy (PEEM), in comparison to scanning techniques such as scanning transmission X-ray microscopy (STXM) and scanning photoemission microscopy (SPEM) that provide spatial information of elemental forms. 

Prepare a 2-pg/mL solution of ribosomal RNA standard provided in TE. Prepare a series of standard RNA concentrations from 0 to 2 pg/mL. Mix equal volumes (100 pL) of diluted RiboGreen Reagent and standard RNAs. Measure the sample fluorescence using a TD-700 fluorometer (excitation approx 480 nm, emission approx 520 nm) and prepare the standard curve. 

Raman spectroscopy in the 1960s. The implementation of FT spectrometers and lasers with output wavelengths that minimized sample fluorescence increased the utility of the technique and fostered application to explosive analysis. 

---