Laser dyes fluorescein

Various xanthene derivatives, including fluorescein, are also used as laser dyes to cover the spectral region from 500 to 700 nm. A modern example is the julolidine-based dye Rhodamine 101 (6.201), which absorbs at 576 nm and lases at 648 nm [79]. 

Further functionalization of mesoporous films with the pH-sensitive dye fluorescein was accomplished by Wirnsberger et al.204 The organosilane used for the actual co-condensation reaction was first prepared by reaction of fluorescein isothiocyanate (FITC) with APTES. The possible use of the dye-modified films as pH sensors was investigated by measurement of the fluorescence after excitation with an Argon laser (488 nm) a dramatic change in fluorescence intensity was observed around pH 8 with a response time of a few seconds. 

Fluorescein structural formula and molecular model. This strongly fluorescent dye has many applications. It is widely used to study retinal circulation and various diseases involving the retina. The technique is known as fluoi escein angiography. Fluorescein can be bound to DNA and other proteins and its fluorescence used as a probe of these molecules and their interactions. Fluorescein is also used for water tracing to provide information on the contamination of underground wells. In addition, it has been used as a laser dye. 

Phthalide derivatives are of major importance in the dye industry, particularly in the area of recording material color formers. Pressure-sensitive carbonless copy paper and thermal recording paper are typical applications. Five principal structural classes have been developed extensively the xanthene dyes (fluorans (208)), 3,3-diarylphthalides (209), spirofluorenes (210), 3,3-bis(di-arylethylene)phthalides (211), and 3-substituted phthalides <84Mi 208-03>. Some of these structures are common to many familiar acid/base indicators, dyes, biological stains, or laser dyes such as fluorescein (212) and phenolphthalein (213) ,... 

Intersystem crossing will obviously be accelerated by heavy-atom sub-stitutients, and no efficient laser dye contains them. The four bromine atoms in eosin (Fig. 9.7) increase its - ISC yield to 76%, as compared to the 3% value observed in fluorescein. Solvents containing heavy atoms (e.g., CBr4) also contribute to buildup in laser dyes, and are avoided. 

Preparation of a series of phycobiliprotein tandem dyes allows multiplexed analysis of different targets in a sample. In addition, since RPE can be excited by the argon-ion laser at 488 nm, a fluorescein-labeled probe can be used concurrently with RPE alone and RPE-tandem conjugates to create a multiplexed system of different fluorescent probes that can be used simultaneously. Table 9.3 shows the different combinations of dyes that can be used in this type of assay with RPE and APC. 

The first, and still the most commonly used, of the tunable lasers were those based upon solutions of organic dyes. The first dye laser was developed by Sorokin and Lankard 05), and used a "chloro-aluminum phthalocyanine" (sic) solution. Tunable dye lasers operating throughout the visible spectrum were soon produced, using dyes such as coumarins, fluorescein, rhodamines, etc. Each dye will emit laser radiation which is continuously tunable over approximately the fluorescence wavelength range of the dye. 

Fig. 3 (a) Normalized excitation and emission spectra of 5-(and-6)-carboxy-fluorescein, succini-midyl ester, rhodamine 6G (R6G), and 6-carboxy-X-rhodamine dyes in pH 7.4 phosphate buffer, (b) Confocal fluorescence image of a mixture of five types of microsphere-DDSN complexes under 488-nm Argon-ion laser excitation. Reproduced with permission from Ref. [12]... 

In the literature, fluorescence spectroscopy in OFD has been limited to the use of ultraviolet (UV) or visible dyes as molecular probes.(1) The most common fluorescent dye used in OFD is fluorescein and its derivatives/21 23) Fluorescein possesses a good fluorescence quantum yield and is commercially available with an isothiocyanate functionality for linking to the polymeric support/24-26 Additionally, selective laser excitation can be performed because the absorbance maximum of fluorescein coincides with the 499-nm laser line emitted from an argon laser. Unfortunately, argon lasers are costly and bulky, thus limiting the practicality of their use. Similar difficulties exist with other popular commercial dyes. 

A fluorescence-activated cell sorter (FACS) is a flow cytometry instrument used to separate and identify cells in a heterogeneous population. Cell mixtures to be sorted are first bound to fluorescent dyes such as fluorescein or phycoerythrin. The labeled cells are then pumped through the instrument and are excited by a laser beam. Cells that fluoresce are detected, and an electrostatic charge is applied. The charged cells are separated using voltage deflection. 

Figure 3 Gel-state versus liquid-state application of vesicles prepared from dilauryl-phosphatidylcholine and septaoxyethylene alkylethers or distearylphosphatidylcholine and cholesterol hemisuccinate. A cross-section of rat skin visualized with confocal laser scanning microscopy after 6 h application. The dye used was fluorescein-phosphatidyl ethanol amine. The vesicles were applied onto rat skin in vivo.

By far most of the work discussed in this review has been based on LIF detection, usually with an 488 nm Ar-ion laser as the excitation source. Only very few other examples exist in the literature where other detection principles were investigated. One of these exceptions is an integrated detection cell for chip CE that has been described by Liang et al. [78]. In combination with the U-shaped separation channel, two additional well aligned channels to take up the excitation and collection fibers where micromachined in a glass plate. The U-cell provides a longitudinal path of 120 -140 pm in length parallel to the flow direction and can be used both for absorption and fluorescence measurements. The absorption detection limit was 0.003 AU ( 6 pM of a fluorescein dye) in the fluorescence mode a detection limit of 3 nM fluorescein (20 000 molecules) was achieved. 

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