Fluorescent Silica Particles

Figure C2.6.2. CSLM image of near hard-sphere silica particles of diameter d = 1050 nm witli a fluorescent core of diameter 400 nm, showing fee stacking (top), hep stacking (bottom middle) and amoriDhous areas (image size 16.3 pm X 16.3 pm, courtesy of Professor A van Blaaderen)...

Tleugabulova, D., Duft, A.M., Zhang, Z., Chen, Y., Brook, M.A. and Brennan, J.D. (2004) Evaluating formation and growth mechanisms of silica particles using fluorescence anisotropy decay analysis. Langmuir, 20, 5924—5932. 

Silica particles have been exploited in virtually every assay or detection strategy that polymer particles have been used in for bioapplication purposes. Recently, fluorescent dye-doped silica nanoparticles have been developed by a number of groups that have similar fluorescence characteristics to quantum dot nanocrystals (Chapter 9, Section 10). Fluorescent silica nanoparticles can be synthesized less expensively than quantum dots due to the fact that the silica particles incorporate standard organic dyes (Ow et al., 2005 Wang et al., 2006) and are not dependent on making reproducible populations of semiconductor particles with precise diameters to tune emission wavelengths. 

Fluorescent silica nanoparticles, called FloDots, were created by Yao et al. (2006) by two synthetic routes. Hydrophilic particles were produced using a reverse micro-emulsion process, wherein detergent micelles formed in a water-in-oil system form discrete nanodroplets in which the silica particles are formed. The addition of water-soluble fluorescent dyes resulted in the entrapment of dye molecules in the silica nanoparticle. In an alternative method, dye molecules were entrapped in silica using the Stober process, which typically results in hydrophobic particles. Either process resulted in luminescent particles that then can be surface modified with... 

Figure 14.23 Silica nanoparticles containing fluorescent dye molecules can be prepared using a reverse micelle suspension process (a) The water-in-oil emulsion is formed with the aqueous phase droplets containing TEOS and dye molecules in detergent, (b) The final particles contain entrapped dye within the silica particle matrix, creating highly fluorescent particles.

Ow et al. (2005) developed an improved method of incorporating fluorescent molecules into silica particles using a modified Stober synthesis, which resulted in both enhanced fluorescence and photostability of the encapsulated dyes. In this two-stage procedure, reactive organic dyes... 

In the preparation of 15 nm core-shell fluorescent silica particles, Ow et al. (2004) reported that the naked core (2.2 nm) alone produced a fluorescence intensity of less than the free dye in solution, presumably due to dye quenching. However, upon addition of the outer silica shell around the core, the brightness of the particles increased to 30 times that of the free dye (using tetramethylrhodamine-5-(and 6)-isothiocyanate (TRITC)). They speculate that shell may protect the core from solvent effects, as evidenced by a lack of spectral shift upon changing the solvent in which the particles are suspended. 

Surface functionalization of silica particles or fluorescent silica particles typically is done using functional alkyl silanes. The process may be used to add a reactive group to the surface of the particles for spontaneous coupling to biomolecules or it may be used to add the appropriate nucleophilic group to the surface, such as an amine or a carboxylate. Silane modification chemistry is discussed in more detail in Chapter 13. 

Add 32 mg of silica nanoparticles (fluorescent or plain) to 20 ml of 1 mM acetic acid containing 1 percent trimethoxysilyl-propyldiethylenetriamine with stirring. Other concentrations of silane derivatives used for particle modification typically range from 1 to 5... 

Nyffenegger, R., Quellet, C., and Rieka, J. (1993) Synthesis of fluorescent, monodisperse, colloidal silica particles./. Colloid Interface Sci. 159, 150-157. 

Yang HH, Qu HY, Lin P, Li SH, Ding MT, Xu JG (2003) Nanometer fluorescent hybrid silica particle as ultrasensitive and photostable biological labels. Analyst 128 462-466... 

Gabriel, U., Charlet, L., Schlapfer, C. W., Vial, J. C., Brachmann, A. Geipel, G. 2001. Uranyl surface speciation on silica particles studied by time-resolved laser-induced fluorescence spectroscopy. Journal of Colloid and Interface Science, 239, 358-368. 

If the nanodispersed species have sizes of the order of 10 nm, diagnostic agents encapsulated in such nanoparticles could potentially cross into human cells. For example, 10 nm diameter, silica-coated, cadmium selenide crystals have been able to transfer into vesicles and be transported by them [901], These protein-sized particles fluoresce for long periods of time making them potentially useful for diagnostic labelling. 

The power of fluorescence detection was illustrated on the separation of PAH s by Yan et al. [64]. The 16 U.S. EPA priority PAHs were separated in isocratic mode in less then 10 min using 100 pm I.D. columns packed with 1.5 pm nonporous octadecyl silica particles. Separation efficiencies of 750,000 plates/m were obtained when the PAHs were detected by ICFD while 300,000-400,000 plates/m were found for OCFD. 

Spectroscopy. It has been shown previoiasly L that the fluorescence spectrum of Ruthenim tris-bipyridine, RuII, is solvent dependent, showing a red shift with increasing solvent polarity. The fluorescence spectrvnn of RuII on silica particles is identicel to that of the excited molecule in water. It will be shown subsequently that the RuII is essentially all botmd to the silica particle, hence the data show that the environment of a probe molec ile such as RuII on silica particles is very polar and similar to water. 

Substantial efforts have been devoted to the development of molecular sensors for dopamine. Raymo et al.70 reported a two-step procedure to coat silica particles with fluorescent 2,7-diazapyrenium dications sensing toward dopamine. The analysis of the fluorescence decay with multiple-equilibria binding model revealed that the electron deficient dications and the electron-rich analytes form 1 1 and 1 2 complexes at the particle/water interface. The interfacial dissociation constants of the 1 1 complexes were 5.6mM and 3.6mM for dopamine and catechol, respectively. Dopamine was dominated by the interaction of its electron-rich dioxyarene fragment with the electron-deficient fluorophore in neutral aqueous environments. Ahn et al.71 reported tripodal oxazoline-based artificial receptors, capable of providing a preorganized hydrophobic environment by rational design, which mimics a hydrophobic pocket predicted for a human D2 receptor. A moderate binding affinity, a dissociation constant of 8.2 mM was obtained by NMR titrations of tripodal oxazoline-based artificial receptor with dopamine in a phosphate buffer solution (pH 7.0). Structurally related ammonium ions, norepinephrine, 2-phenylethylamine,... 

Figure 1.49 3D confocal fluorescent micrograph of silica particles in low density polyethylene. (Reproduced with permission from A.R. Clarke and C.N. Eberhardt, Microscopy Techniques for Materials Science, Woodhead Publishing Ltd, Cambridge UK. 2002 Woodhead Publishing Ltd.)... 

Figure 1.49 shows an example of using confocal fluorescent microscopy to reveal microscopic features in a specimen. The specimen is low density polyethylene (LDPE) containing fluores-cently labeled silica particles. The particle size and distribution in the polymer matrix can be clearly revealed by 3D confocal microscopy. Thus, confocal microscopy provides us a new dimension in light microscopy for materials characterization, even though its applications in materials science are not as broad as in biology. 

An integrated pTAS system for the detection of bacteria including lysis, DNA purification, PCR and fluorescence readout has also been published recently [113]. A microfluidic plastic chip with integrated porous pol5mier monoliths and silica particles for lysis and nucleic acid isolation was used for detection (Fig. 8). A custom-made base device provided liquid actuation and off-chip valving by stopping liquid flow from the exits of the chip, utilizing the incompressibility of liquids. Detection of 1.25 x 10 cells of B. subtilis was demonstrated with all assay steps performed on-chip. 

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