Introduction

Fluorescent particles are prepared by incorporating selected fluorophores into monodisperse silica particles through swelling process or copolymerizing silica with various organic fluorescent dyes, which produce fluorophores labeled silica particles with satisfactory properties. Functional groups or biological macromolecules (carboxyl, amino, streptavidin, biotin, etc.) can be quantitatively modified on the surface of microsphere as linking groups for immunoanalysis, making fluorescent silica microspheres have a wide range of applications, including lateral chromatography, cell imaging, microfluidics and fluorescence enzyme-related immunosorbent assay. Fluorescent particles with single or multiple fluorophores are available in various sizes, emission spectra and combinations.    Many are suitable for uses in flow cytometry, fluorescence microscopy, phagocytosis studies, and cell labeling[1-4].

Abvigen Inc. offers a wide range of Fluorescent Silica Particles, including Green Silica Fluorescent Particles, Blue Silica Fluorescent Particles, Red Silica Fluorescent Particles and Orange Silica Fluorescent Particles. The product size is adjustable within the range of nanometers to micrometers, and can be further flexibly adjusted according to customer requirements and use conditions to achieve customized supply.

Preparation Method

One-step self-assembly process[5]: To synthesize mesoporous silica particles, the Origami type of synthesis was used with the addition of fluorescent lasing dyes. The materials used were tetraethylorthosilicate (TEOS，99.99+%，Aldrich)，cetyltrimethylammoniumchloride (CTACI, 25 wt.% aqueous solution, Pflatz & Bauer), formamide (99%, Aldrich), hydrochloric acid (37.6 wt.% aqueous solution, SafeCote), and rhodamine 6Gperchlorate dye (Sigma-Aldrich). All chemicals were used as received. Synthesis: The surfactant, acid, dye, formamide, and distilled water (Coming, AG-1b, 1MΩ cm) were stirred in a polypropylene bottle at room temperature for 2 h, after which TEOS was added and the solution stirred for approximately 5 min. The solution was then kept under quiescent conditions for 3 days. The molar ratio of H2O: HCI: formamide: CTACI: TEOS: R6G was 100:7.8:9.5:0.11:0.13:6x10-4-0.01. The materials so formed were filtered, washed with copious amounts of water, and air dried.

Templated sol-gel self assembly[6]: The synthesis of ultrabright fluorescent mesoporous silica nanoparticles (UFSNPs) of various sizes loaded with different amounts of fluorescent dye (Rhodamine 6G) is reported here. The dye is physically entrapped inside the nanochannels of the silica matrix created during templated sol-gel self assembly. Due to the specific nanoenvironment, the fluorescence of the encapsulated dye molecules remains unquenched up to very high concentrations, which results in relatively high fluorescence. The particle size (ranging from 20-50 nm) and dye loading (0.8-9.3 mg dye per g particles) are controlled by the timing of the synthesis and the concentration of several organotriethoxysilanes, which are coprecursors of silica. The quantum yields of the encapsulated dye range from 0.65 to 1.0. The relative brightness of a single particle is equivalent to the fluorescence of 30-770 free nondimerized R6G dye molecules in water, or to that of 1.5-39 CdSe/ZnS quantum dots. Despite the presence of some hydrophobic groups on the particles' surfaces, colloidal suspensions of the particles are relatively stable (as monitored for 120 days).

Functionalized by incorporating a covalently labeled fluorescent dye[7]: Chemically modified silica particles were synthesized using hydrolysis and condensation reactions via the St?ber process. These silica particles were subsequently functionalized by incorporating a covalently labeled fluorescent dye. The synthesized silica particles were dyed with rhodamine B isothiocyanate (RBITC) using the following procedure. RBITC solution (1 × 10?4 M) was prepared in dimethylformamide (DMF); 20 L of this solution was diluted again into 100 L DMF. This diluted RBITC solution was then added to the synthesized silica particles, and the mixture was stirred for 2 h in the dark. Finally, the products were washed with DMF followed by ethanol to remove any residual RBITC. The synthesized fluorescent silica particles were stored in a brown-colored bottle at 4℃. When used in the experiments, the particles were dispersed in DI water and sonicated for maximum dispersion.

Preparation of dually fluorescent silica nanoparticles[8]: Two cationic dyes (9-aminoacridine and safranine) were incorporated in silica nanoparticles prepared by a modified Stober method, with the aim of investigating fluorescence properties and intra-particle energy transfer.

Synthesis of fluorescent, monodisperse, colloidal silica particles[9]: The preparation of colloidal, fluorescently labeled silica particles by controlled hydrolysis and polycondensation of tetramethoxysilane (TMOS) in the presence of a silylated fluorescein derivative is described. Monodisperse particles with average radii of 80 nm and a narrow size distribution with a standard deviation lower than 8% are obtained using the seeded growth technique, whereas homogeneous nucleation yields particles with a bimodal size distribution. The apparent degree of labeling ranges from 5 × 10-8 to 2 × 10-6 mol equivalent of fluorescein per gram of silica, depending on the labeling procedure.

Covalent modification-reverse micelle technique[10]: Nanometer-sized fluorescent hybrid silica (NFHS) particles were prepared for use as sensitive and photostable fluorescent probes in biological staining and diagnostics. The first step of the synthesis involves the covalent modification of 3-aminopropyltrimethoxysilane with an organic fluorophore, such as fluorescein isothiocyanate, under N2 atmosphere for getting a fluorescent silica precursor. Then the NFHS particles, with a diameter of well below 40 nm, were prepared by controlled hydrolysis of the fluorescent silica precursor with tetramethoxysilane (TMOS) using the reverse micelle technique. The fluorophores are dispersed homogeneously in the silica network of the NFHS particles and well protected from the environmental oxygen. Furthermore, since the fluorophores are covalently bound to the silica network, there is no migration, aggregation and leakage of the fluorophores. In comparison with common single organic fluorophores, these particle probes are brighter, more stable against photobleaching and do not suffer from intermittent on/off light emission (blinking). We have used these newly developed NFHS particles as a fluorescent marker to label antibodies, using silica immobilization method, for the immunoassay of human α-fetoprotein (AFP). The detection limit of this method was down to 0.05 ng mL?1 under our current experimental conditions.

Application

Phagocytosis studies;

Cell imaging;

Tracking;

Calibration of flow cytometry;

Lateral chromatography;

Microfluidics;

Fluorescence enzyme-related immunosorbent assay;

Fluorescence microscopy

Advantages

Monodisperse Particles;

High particle size uniformity;

High-intensity fluorescence;

Available with red, green orange and blue fluorescence;

Designed with carboxylic acid groups (COOH) and amino groups (NH2) on the particle surface for the covalent binding of proteins, antibodies or other molecules;

Offered with streptavidin on the surface for binding of biotinylated molecules;

Outstanding long-term stability under proper storage conditions (both color and fluorescence);

Spherical shape;

Great resistance to photobleaching;

Minimized dye leaching