Optimizing the performance of silica nanoparticles functionalized with a near-infrared fluorescent dye for bioimaging applications

Abstract

Modified fluorescent nanoparticles continue to emerge as promising candidates for drug delivery, bioimaging, and labeling tools for various biomedical applications. The ability of nanomaterials to fluorescently label cells allow for the enhanced detection and understanding of diseases. Silica nanoparticles have a variety of unique properties that can be harnessed for many different applications, causing their increased popularity. In combination with an organic dye, fluorescent nanoparticles demonstrate a vast range of advantageous properties including long photostability, surface modification, and signal amplification, thus allowing ease of manipulation to best suit bioimaging purposes. In this study, the Stöber method with tetraethyl orthosilicate (TEOS) and a fluorescent dye sulfo-Cy5-amine was used to synthesize fluorescent silica nanoparticles. The fluorescence spectra, zeta potential, quantum yield, cytotoxicity, and photostability were evaluated. The increased intracellular uptake and photostability of the dye-silica nanoparticles show their potential for bioimaging.

Keywords: fluorescence imaging; photostability; silica nanoparticles; stöber method.

Figure 1.

Absorbance spectra of different dye-loaded sCy5-SiNPs (μg of dye per mg of silica) (A) and the absorbance of samples with varying dye content at 650 nm (the dotted line represents the best linear fit) (B).

Figure 2.

Fluorescence spectra of dye-loaded sCy5-SiNPs (μg of dye per mg of silica) (A) and their fluorescence intensity at 660 nm (B).

Figure 3.

Relative quantum yield values of sCy5 encapsulated nanoparticles.

Figure 4.

Transmission electron microscopy (TEM) images of bare and dye-incorporated silica nanoparticles prepared with different content of sCy5 dye: (A) bare; (B) low-NP; (C) high-NP.

Figure 5.

Percent degradation for each SiNP sample type over a period of 60 minutes of irradiation at 694 nm.

Figure 6.

Cellular internalization of NP-high (A), NP-low (B), bare SiNP (C), and media-only control (D) after 4 hours of incubation. Cytotoxicity of SiNP samples on MDA-MB-231 breast cancer cell line (E). Photostability of internalized SiNP samples (n=3, F). Blue color corresponds to the DAPI channel, and red color indicates fluorescence in the Cy-5 channel.

Scheme 1:

Synthesis of bare nanoparticles via Stöber method: (a) hydrolysis; (b) condensation. Reaction conditions: 10 mM chloride-ammonium hydroxide buffer, pH 9.0, cyclohexane, 50°C, 24h.

Scheme 2.

Synthesis of cyanine dye-incorporated silica nanoparticles. Conditions: (a) conjugation: sCy5-NHS ester was reacted with APTES at a molar ratio of 1.2:1 (APTES:dye). Reaction proceeded at RT for 24 h in dark; (b) first reaction of hydrolysis and condensation: 10 mM chloride-ammonium hydroxide buffer, pH 9.0, 50^oC, 30 min; (c) second reaction of hydrolysis/condensation and stabilization reactions: TEOS, cyclohexane, 50^oC, 24 h.