Hybrid silica nanoparticles traceable by fluorescence and FT-IR spectroscopy: Preparation, characterization and preliminary biological studies

The novel reporters, unimodal FT-IR-traceable, bimodal fluorescent and FT-IR-traceable hybrid silica (SiO₂) nanoparticles (NPs), were prepared using the Stöber methodology. Initially, the basic Stöber co-hydrolysis of tetraethoxysilane (TEOS) and of the iron-complexed alkyl-triethoxysilane (EtO)₃Si-l-(η⁴-2E,4E-dienyl)- Fe(CO)₃(0) (L = alkyl linker, DIT-tag reagent 3) afforded unimodal 13.7 ± 2 nm-sized SiO₂@DIT-tag₆₀ NPs. These NPs can incorporate an organometallic FT-IR sensitive (η⁴-2E,4E-dienyl)- tricarbonyliron(0) complex moiety which acts as a sensitive FT-IR traceable species due to the strong iron complex ν_FeCO vibrational bands that appear in the 1996-2063 cm^-1 region, a region free of any parasitic band. The sensitivity of the FT-IR-based detection of SiO ₂@DIT-tag₆₀ NPs has been determined using incremental mixtures of B16 melanoma cell lysates as a biological medium model. The detection limit was found to be 0.190 μg of Fe per mg of B16 cell lysate. In the second step, bimodal hybrid fluorescent and FT-IR-traceable 35.7 ± 5 nm sized SiO₂@DIT-tag₂₀@FITC NPs were similarly fabricated, in order to enable both fluorescence and FT-IR spectroscopy detection. This has been readily obtained through the straightforward co-incorporation of an additional fluorescein-containing alkyl-triethoxysilane conjugate FITC-APTES (FITC: fluorescein isothiocyanate, APTES: (3-aminopropyl)triethoxysilane). Subsequent surface modification with APTES of the resulting SiO₂@DIT-tag₂₀@FITC NPs afforded amine functionalized 34.4 ± 6 nm-sized SiO₂@DIT-tag ₂₀@FITC@NH₂ NPs that were readily endocytosed by B16 melanoma cells. All these novel hybrid silica NPs have been fully characterized by FT-IR spectroscopy, high resolution TEM/SEM (HR-TEM/SEM) with elemental energy-dispersive X-ray spectroscopy (compositional EDAX analysis), dynamic light scattering (DLS), ζ potential measurements, and inductively coupled plasma-optical emission spectroscopy (ICP-OES). Preliminary biological studies demonstrated the non-toxicity of the NPs. No observable modification in the B16 cells' morphology or mortality was seen after the internalization of SiO ₂@DIT-tag₂₀@FITC@NH₂ NPs.