1 Versatile Synthesis Strategy for Carboxylic Acid-functionalized Upconverting Nanophosphors as Biological Labels Zhigang Chen, Huili Chen, He Hu, Mengxiao Yu, Fuyou Li,* Qiang Zhang,Zhiguo Zhou, Tao Yi, and Chunhui Huang* J. AM. CHEM. SOC. 2008, 130, 演講者：洪柏楷

2 Outline Phosphors Synthesis of Oleic Acid-Capped UCNPs Converting Hydrophobic UCNPs into Hydrophilic Preparation of Streptavidin-Functionalized UCNPs Detection of DNA

3 Phosphors Downconverting phosphors Upconverting phosphors

4 Downconversion fluorescent materials 1. Organic dyes 2. Semiconductor nanocrystal

5 Up-converting rare-earth nanophosphors (UCNPs) 1. sharp absorption and emission lines 2. high quantum yields 3. long lifetimes 4. photostability

6 oleic acid 2,6-diaminohexanoic acid (Lysine) 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC)

7 Hydroxy-2,5-dioxopyrrolidine-3- sulfonicacid sodium salt (Sulfo-NHS) 4-Morpholineethanesulfonic acid (MES) 2-Amino-2-hydroxymethyl- propane-1,3-diol (Tris)

8 Synthesis of Oleic Acid-Capped UCNPs Ln: 78 mol% Y + 20 mol%Yb + 2 mol%Er NaOH + water + ethanol + oleic acid rare-earth chloride (LnCl 3 ) transfer to autoclave 160 °C 8 h cyclohexane + ethanol oleic acid-capped UCNPs 1.0 M NaF

9 NaYF 4 : 20 mol%Yb, 2 mol % Er

10 Converting Hydrophobic UCNPs into Hydrophilic UCNP + cyclohexane + tert-butanol + water 5 wt % K 2 CO 3 aqueous solution KMnO 4 + NaIO 4 (Lemieux-von Rudloff reagent) 40 °C 48 h. wash with deionized water acetone ethanol. HCl (50 mL) pH min washed twice with deionized water hydrophilic UCNPs

11 Oxidized NaYF 4 :Yb,Er

12 TEM oxidized NaYF 4 :Yb,ErNaYF 4 :Yb,Er

13 TEM NaYF 4 :Yb, Tmoxidized NaYF 4 :Yb, Tm

14 TEM NaYF 4 :Yb, Hooxidized NaYF 4 :Yb, Ho

15 EDXA spectra NaYF 4 :Yb,Er oxidized NaYF 4 :Yb,Er

16 XRD pattern * cubic phase ▼ hexagonal phase oxidized NaYF 4 :Yb,Er NaYF 4 :Yb,Er

17 1 H NMR Spectra undoped NaYF 4 oxidized NaYF 4

18 FTIR spectra NaYF 4 :Yb,Er oxidized NaYF 4 :Yb,Er

19 Thermogravimetric analysis (TGA) oxidized NaYF 4 :Yb,Er NaYF 4 :Yb,Er % 7.55 %

20 Oxidized NaYF 4 :Yb,Er water DMF DMSO

21 Dynamic light scattering analysis

22 Luminescence continuous-wave excitation at 980 nm red filtergreen filter

23 Luminescence spectra (A) NaYF 4 :Yb,Er (B) Oxidized NaYF 4 :Yb,Er 2 H 11/2 to 4 I 15/2 4 S 3/2 to 4 I 15/2 4 F 9/2 to 4 I 15/2 ( I 540 /I 654 ) decrease

24 Luminescence spectra

25 Energy level diagram Leyu Wang ; Yadong Li Chem. Mater. 2007, 19,

26 Luminescence spectra

27 Preparation of Streptavidin- Functionalized UCNPs oxidized NaYF 4 :Yb,Er + EDC + sulfo- NHS + MES buffer 8 h water PBS buffer solution containing streptavidin 4 °C 48 h Lysine water Streptavidin-Functionalized UCNPs

28 Streptavidin-Functionalized UCNPs

29 Oligonucleotide 5 ' - (biotin)-GATGAGTATTGATGC-3 ' (as Capture-DNA) 5 ' -CGAATAGTTCCATTG-(TAMRA)-3 ' (as Report-DNA) 5 ' -CAATGGAACTATTCG GCATCAATACTCATC-3 ' (as Target-DNA)

30 DNA nanosensors based on UCNPs

31 TARMA-labeled reporter DNA absorption emission oxidized NaYF 4 : Yb, Er

32 Detection of DNA streptavidin-functionalized UCNPs + Report-DNA + Capture-DNA + buffered solution. 37 °C for 20 min Target-DNA 980 nm luminescence spectrum

33 Luminescence spectra

34 DNA nanosensors based on UCNPs Fluorescence resonant energy transfer

35

36 Conclusions We have demonstrated a new, efficient and versatile procedure for converting hydrophobic UCNPs into water-soluble and carboxylic acid- functionalized derivatives by directly oxidizing oleic acid ligands to azelaic acid. This procedure is not limited to hydrophobic UCNPs,and it can easily be applied to other hydrophobic nanoparticles (including rare earth, semiconductor and metal nanoparticles) where only surface ligands can be oxidized. The presence of free carboxylic acid groups on the surface of azelaic acid- capped UCNPs allows further conjugation with various biomolecules, and streptavidin-functionalized UCNPs derived from azelaic acid-capped precursors provides a novel approach for detecting and/or binding to a broad range of biotinylated proteins or antibodies.