Syntese af Cd-fri InP / ZnS Quantum Dots Velegnet til Biomedical Applications
Summary
In this protocol, the synthesis of Cd-free InP/ZnS quantum dots (QDs) is detailed. InP-based QDs are gaining popularity due to the toxicity of Cd2+ ions that may be released through nanoparticle degradation. After synthesis, QDs are solubilized in water using an amphiphilic polymer for use in biomedical applications.
Abstract
Fluorescent nanocrystals, specifically quantum dots, have been a useful tool for many biomedical applications. For successful use in biological systems, quantum dots should be highly fluorescent and small/monodisperse in size. While commonly used cadmium-based quantum dots possess these qualities, they are potentially toxic due to the possible release of Cd2+ ions through nanoparticle degradation. Indium-based quantum dots, specifically InP/ZnS, have recently been explored as a viable alternative to cadmium-based quantum dots due to their relatively similar fluorescence characteristics and size. The synthesis presented here uses standard hot-injection techniques for effective nanoparticle growth; however, nanoparticle properties such as size, emission wavelength, and emission intensity can drastically change due to small changes in the reaction conditions. Therefore, reaction conditions such temperature, reaction duration, and precursor concentration should be maintained precisely to yield reproducible products. Because quantum dots are not inherently soluble in aqueous solutions, they must also undergo surface modification to impart solubility in water. In this protocol, an amphiphilic polymer is used to interact with both hydrophobic ligands on the quantum dot surface and bulk solvent water molecules. Here, a detailed protocol is provided for the synthesis of highly fluorescent InP/ZnS quantum dots that are suitable for use in biomedical applications.
Introduction
Quantum prikker (QDs) er halvledende nanokrystaller, der udviser fluorescerende egenskaber, når bestråles med lys 1. På grund af deres lille størrelse (2-5 nm), som er magen til mange større biomolekyler, og let biofunctionalization, QDs er et særdeles attraktivt middel til biomedicinske anvendelser. De har fundet anvendelse i biologisk mærkning, single-molekyle live-cell imaging, drug delivery, in vivo imaging, påvisning af patogener, og celle sporing, blandt mange andre anvendelser 2-8.
Cd-baserede QDs er mest almindeligt anvendt i biomedicinske anvendelser på grund af deres intense fluorescens og smalle peak emission bredder 9. Imidlertid er der rejst spørgsmål på grund af mulige toksicitet af Cd 2+ ioner 10, som kan frigives ved nedbrydning af nanopartikel. For nylig har InP-baserede QDs blevet udforsket som et alternativ til Cd-baserede QDs fordi de opretholder mange fluorescens egenskaberaf Cd-baserede QDs og kan være mere biokompatible 11. CD-baserede QDs har vist sig at være signifikant mere toksiske end InP-baserede QDs i in vitro-assays ved koncentrationer så lave som 10 pM efter kun 48 timer 11.
Fluorescensemissionen farve QDs er størrelse-afstemmelige 1. Det er, som størrelsen af QD stiger, fluorescensemissionen er rødforskudt. Størrelsen og størrelse dispersitet QD produkter kan ændres ved at ændre temperaturen, reaktionen varighed eller precursor koncentration betingelser under reaktionen 12. Mens toppen udledning af InP QDs er typisk bredere og mindre intens end Cd-baserede QDs kan InP QDs ske i en lang række farver designet til at undgå spektral overlapning, og er tilstrækkeligt intens til de fleste biomedicinske anvendelser 12. Syntesen beskrevet i denne protokol giver QDs med en rød top emission centreret ved 600 nm.
Flere trin er taget ofter syntese af QD kerner til at opretholde den optiske integritet QDs og gøre dem kompatible til biologiske anvendelser. Overfladen af QD kerne skal beskyttes mod oxidation eller overfladefejl, der kan forårsage afkølende; Derfor er en ZnS shell coatet over kernen til at producere InP / ZnS (kerne / skal) QDs 13. Denne belægning har vist sig at beskytte fotoluminescens af QD produkt. Tilstedeværelsen af zinkioner under InP QD syntese er blevet vist at begrænse overflade defekter, samt nedgang størrelsesfordeling 12. Selv med tilstedeværelsen af Zn2 + i reaktionsmediet, syntese af InZnP er højst usandsynligt 12. Efter coating er resulterende InP / ZnS QDs belagt med hydrofobe ligander, såsom trioctylphosphinoxid (TOPO) eller oleylamin 12,14. En amfifil polymer kan interagere med hydrofobe ligander på QD overflade samt bulk-vandmolekyler at bibringe vandopløselighed 15. Amfifile polymerer med carboxylate kemiske grupper kan anvendes som "kemiske håndtag" for yderligere at funktionalisere de QDs.
Denne protokol beskriver syntesen og funktionalisering af vandopløselige InP / ZnS QDs med meget intens fluorescens emission og relativt lille størrelse-dispersitet. Disse QDs er potentielt mindre toksiske end almindeligt anvendte CdSe / ZnS QDs. Heri syntesen af InP / ZnS QDs giver et praktisk alternativ til Cd-baserede QDs til biomedicinske anvendelser.
Protocol
Representative Results
Discussion
Denne protokol detaljer syntesen af stærkt fluorescerende InP / ZnS QDs, der kan bruges i mange biologiske systemer. QD produkter syntetiserede her udviste en enkelt top fluorescensemission centreret ved 600 nm med en FWHM på 73 nm (figur 1), som er sammenlignelig med andre tidligere beskrevne synteser 12. Reaktionstid og reaktionstemperatur er ekstremt afgørende skridt på grund af deres dybtgående effekt på QD syntese kvalitet og gentagelsesnøjagtighed. Efter opløsning i vand, …
Divulgazioni
The authors have nothing to disclose.
Acknowledgements
Forfatterne takker Kemisk Institut og Graduate College på Missouri State University for deres støtte til dette projekt. Vi anerkender også Electron Microscopy Laboratory ved Frederick Nationallaboratoriet for Cancer Research for brug af deres transmissions elektron mikroskop og kulstof-coatede gitre.
Materials
| Oleylamine | Acros | 129540010 | |
| Zinc (II) chloride | Sigma | 030-003-00-2 | |
| Indium (III) chloride | Chem-Impex | 24560 | |
| Tris(dimethylamino)phosphine | Encompass | 50-901-10500 | |
| 1-dodecanethiol | Acros | 117625000 | |
| Hexanes | Fisher Sci | H292-4 | |
| Acetone | TransChemical | UN 1090 | |
| Zinc Stearate | Aldrich Chem | 307564-1KG | |
| Tetrahydrofuran | Acros | 34845-0010 | |
| Molecular Water | Fisher Sci | BP2470-1 | |
| Poly(maleic anhyrdride-alt-1-tetradecene), 3-(dimethylamino)-1-propylamine derivative | Sigma | 90771-1G | |
| Boric acid | Fisher Sci | BP168-500 | |
| Sodium Tetraborate Decahydrate | Fisher Sci | BP175-500 | |
| Rhodamine B | Aldrich Chem | R95-3 | |
| Nitrogen gas | Airgas | UN1066 | |
| Trypan blue | Thermo Sci | SV30084.01 | |
| 3 mL plastic Luer-lock syringe | BD | 309657 | |
| Luer-lock Needle | Air-Tite | 8300014471 | 4 inch, 22 gauge |
| 50 mL polypropyene centrifuge tube | Falcon | 352098 | |
| 250 mL centrifuge bottle | Thermo Sci | 05-562-23 | Nalgene PPCO |
| 5 mL centrifuge tubes | Argos-Tech | T2076 | |
| 1.5 mL microcentrifuge tubes | Bio Plas | 4150 | |
| 0.1 μm Syringe filter | Whatman | 6786-1301 | Puradisc 13 mm nylon filter |
| Slide-A-Lyzer MINI Dialysis Unit | Thermo Sci | 69590 | 20,000 MWCO |
| Rotary Evaporator | Heidolph | ||
| Centrifuge 5072 | Eppendorf | Swinging Bucket with 50 mL tube adapters | |
| Lambda 650 UV/VIS Spectrometer | Perkin Elmer | UV-Vis Spectrophotometer | |
| LS 55 Fluorescence Spectrometer | Perkin Elmer | Fluorometer | |
| Axio Observer.A1 | Zeiss | epifluorescence microscope | |
| AxioCam MRm | Zeiss | CCD Camera | |
| Tecnai TF20 Microscope | FEI | Transmisison Electron Miscroscope | |
| TEM Eagle CCD | FEI | TEM CCD Camera | |
| NanoBrook Omni DLS | Brookhaven | Dynamic Light Scattering Instrument |
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