Keywords

• Fluorescent magnetic nanoparticles;
• Quantum dots;
• Magnetic;
• Silica;
• Flow cytometric analysis

1. Introduction

Multifunctional nanoparticles are attracting more and more attentions of material scientists, chemists and biologists, since such multifunctional particles combine multiple properties and have various applications [1], [2], [3], [4] and [5]. Especially, considerable efforts have been devoted to design fluorescent magnetic nanoparticles (FMNPs) with both fluorescence and magnetism functionalities for potential application as dual-modality imaging probes [6], [7], [8], [9], [10] and [11]. In addition to the use as imaging probes, FMNPs have potential applications in bio-analysis [12] and [13]. FMNPs can be employed to separate and enrich the analytes from complicated samples, and then the enriched analytes can be detected with the fluorescent signal generated by FMNPs. The preparations of FMNPs have been extensively reported these years [14], [15], [16], [17] and [18]. However, one of the common problems during the preparation of FMNPs is fluorescent quenching, resulting in the low QY of FMNPs [15] and [19]. The fluorescent quenching is mainly owing to two factors. The first one is the photo/chemical stability of the used fluorophores. FMNPs are usually prepared and used in the complicated chemical solution, and many chemicals could be the quencher of fluorophores [20], [21], [22], [23] and [24]. The second factor is the influence of MNPs because of the strong absorption cross section of the magnetic nanoparticles [15] and the possible energy transfer between fluorophores and MNPs [25]. The second quenching effect could be weakened by adjusting the mol ratio of embedded fluorophores to MNPs [17]. However, the saturation magnetization of the FMNPs will be insufficient if the ratio of MNPs to fluorophores is too low. Thus, the most promising solution to solve fluorescent quenching is the selection of proper fluorophores.

Semiconductor nanocrystals called QDs [26] have high photostability, high emission quantum yield, narrow emission peaks, size-dependent wavelength tunability in comparison with organic dyes and fluorescent proteins, which make them more interesting for potential biomedical application [27]. Most of QDs are hydrophobic, since they are capped by organic ligands. And they are photo stable in organic solution. However, hydrophobic QDs should be transferred into hydrophile for biomedical application. One of the most widely used modification methods is silanization. However, the fluorescence of QDs will be quenched in various degrees when they are transferred into water phase [28] and [29]. Our group has done some work about the changes of optical property of QDs before and after silica-coating. And study data indicated that QDs with appropriate structure and composition of shells can retain the initial QY more efficiently after silanization [30]. Based on the results from our past work, in this study, the well-designed seven layered shell/core QDs (CdSe/CdS/CdS/Cd_0.75Zn_0.25S/Cd_0.5Zn_0.5S/Cd_0.25Zn_0.75S/ZnS/ZnS) were selected to prepare highly florescent FMNPs with the classical silica-coating via the reverse microemulsion approach [31]. After the successful preparation of FMNPs, the prepared samples were chemically-activated by amino groups for antibody-labeling and the following target-enrichment and subsequent fluorescent detection on a flow cytometry.

2. Materials and methods

2.1. Materials

Selenium powder (100 mesh, 99.99%, Aldrich), cadmium oxide (CdO, 99.5%, Aldrich), zinc oxide (ZnO, 99.9%, Sigma), Fe(acac)₃ (Alfa Aesar), 1,2-hexadecanediol (Alfa Aesar), benzyl ether (Alfa Aesar), sulphur (99.98%, Aldrich), tri-n-butylphosphine (TBP, 90%, TCI, Japan), tri-n-octylphosphine oxide (TOPO, 90%,Aldrich), octadecylamino (ODA, 90%, ACROS), 1-octadecene (ODE, 90%, ACROS), oleic acid (OA, 90%, Aldrich), Carboxyl-polystyrene (PS) microspheres (Tianjin BaseLine ChromTech Research Center); Bovine Serum Albumin (BSA), human IgG: purified total IgG from normal human serum, in which heavy chain is 50,000 Da, light chain is 25,000 Da. Goat anti-human IgG: pure human total IgG immune against goat until 1:100,000 (ELISA), above supplied by BEIJING DINGGUO BIOTECHNOLOGY CO. Ltd. 1-Ethyl-3-(3-dimethyllaminopropyl) carbodiimide hydrochloride (EDC, GL Biochem (Shanghai) Ltd.), 3-aminopropyltriethoxysilane (APTES, Alfa Aesar), NP-40 (Fluka), TEOS, aqueous ammonia solution (25 wt.%), cyclohexane, glutaraldehyde, acetone, and argon (oxygen free) were obtained from local suppliers. All chemicals were used without further purification.