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光在生物檢測及造影的應用

光在生物檢測及造影的應用. 台灣大學化學系 陳昭岑. H. C. N. C. H. 3. N. N. H. M. a. u. v. e. i. n. e. A. l. i. z. a. r. i. n. 3. H. C. 3. H. N. +. 2. H. C. N. C. H. C. H. 3. 3. 3. H. N. N. N. +. 2. H. 化工之父. William Perkin. 1856~1906. VIOLET. ORANGE. RED. YELLOW.

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光在生物檢測及造影的應用

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  1. 光在生物檢測及造影的應用 台灣大學化學系陳昭岑

  2. H C N C H 3 N N H M a u v e i n e A l i z a r i n 3 H C 3 H N + 2 H C N C H C H 3 3 3 H N N N + 2 H 化工之父 William Perkin 1856~1906

  3. VIOLET ORANGE RED YELLOW BLUE GREEN Electromagnetic Spectrum Increasing ν 1019Hz 1015Hz 1013Hz 0.1 nm 200 nm 400 nm 800 nm 2 μm 50 μm Increasing λ

  4. T1 Luminescence Jobloñski Diagram Intersystem Crossing S1 hνA hνF hνP 2 S0 1 0 Absorption Fluorescence Phosphorescence

  5. Complimentary Color Table

  6. De Cola’s White Light Material Absorption Spectrum Blue: Ir Red: Eu Blank: Ir+Eu Fluorescence Spectrum Paolo Coppo, Marco Duati, Valery N. Kozhevnikov, Johannes W. Hofstraat, and Luisa De Cola* Angew. Chem. Int. Ed.2005, 44, 1806–1810

  7. Labeling v.s. Sensing Probes Direct Readout Diagnostic Reagent Powerful Screening Tool Genomics Proteomics Medicine

  8. Labeling v.s. Sensing Probes • Labeling:Photophysical properties remain the same before and after recognition • Sensing:Photophysical properties (wavelength, intensity, polarization, half life) change before and after recognition Interference from the analytes Chemical modifications required

  9. The Flow of Biological Informations Replication DNA DNA (deoxyribonucleic acid) Transcription RNA (ribonucleic acid) Translation Proteins Cell Structures and Functions

  10. Coding strand (sense) 5’ +1 +2 +3…… 3’ 5’ 3’ 3000-4000 proteins in E. coli; 30000-40000 in a human being

  11. Telomere Centramere

  12. - O 5 ' - B a s e - O O P O O 1 ' - - H + O P O H O H O B a s e - H O O H 2 ( n u c l e o t i d e ) H O H H 3 ' H P h o s p h o r i c a c i d H O H H O H H + - O - - O O P O B a s e O 5’ 5 ' B a s e - H O O P O O H 2 H H H O H H H H H O H 3 ' H O B a s e O P O O O H H Backbone: Hydrophilic Region H H 3 ' H O B a s e O O O P H H O H H 3 ' H 3’ O H D N A ( o r R N A ) Formation of Nucleic Acids O

  13. Primer strand

  14. Animation of sanger method

  15. Mask Resist SiO2 layer Silicon wafer Photolithography Process for Producing Integrated Circuit Chips Solvent Washing Dissolve resist Etch SiO2

  16. A A A A A A A G G G A A A A A A A 生物晶片製作

  17. Noncomplementary Complementary ! DNA Microarrays Wash DNA Chip

  18. －G≡C－ Minor Groove H A d e n i n e N H O N A＝T N G≡C N T h y m i n e N H N A＝T N D e o x y r i b o s e O C≡G D e o x y r i b o s e A T T＝A Major Groove H O A＝T H N N G u a n i n e C≡G N C y t o s i n e N N H G≡C N N D e o x y r i b o s e T＝A O N H D e o x y r i b o s e T＝A H G C T＝A Double-Helical Structure of DNA

  19. Base-discriminating fluorescent (BDF) nucleosides abs=315nm em =442nm (=0.118) abs=327nm em =397, 427nm (=0.117)

  20. =0.011 =0.081

  21. Molecular beacons

  22. Energy Transfer Chromophore Complementary ssDNA Noncomplementary ssDNA PNA probe Cationic conjugated polymer Peptide nucleic acid (PNA) probes

  23. Noncomplementary ssDNA Complementary ssDNA dsDNA ssDNA-C* CP Chromophore-Labeled Single-Stranded DNA

  24. Sensing assays

  25. Mechanisms of Detection Method ELISA

  26. Receptor Analyte Indicator The competitively sensing system Differential receptors array

  27. Indicators

  28. J. Am. Chem. Soc. 2005, 127, 4888-4894

  29. J. Am. Chem. Soc. 2005, 127, 4888-4894

  30. H2O2… • A major reactive oxygen species (ROS) and a source of oxidative stress. • Connecting to aging and severe human diseases, ex: cancer, cardiovascular disorder, Alzheimer’s and related neurodegenerative diseases. • A second messenger in cellular signal transduction that affect cell proliferation and cell death. ex: trigger mitogen-activated protein (MAP) kinase and nuclear factor κB (NF-κB) pathways.

  31. (DCFH) Dihydro analogues of fluorescent dyes (DCFH-DA) Ref: Free Rad. Biol. Med.1999, 27, 146-159

  32. Bio/Nano

  33. 奈米粒子種類 奈米粒子 金屬材質 非金屬材質 金屬氧化物 單一金屬 合金 (TiO2……..) (Au, Ag, Pd….) （半導體性質..）

  34. 量子侷限效應(quantum confinement effect) • Density of states • Energy splitting near band-edge • Size-dependent optical properties National Taiwan University Center for Nano Science and Technology http://nanost.ntu.edu.tw台灣大學奈米科技研究中心

  35. Alivisatos and his research team pioneered the use of quantum dots to label cells.

  36. 實驗三 發光性半導體奈米微粒之合成與鑑定(零維度奈米材料製備) 周必泰教授 1.合成CdSe/ZnX (X=S, Se) CdO/TOPO/HDA Se/TBP Zn/X 2.合成出發光性半導體奈米微粒之後以TEM及Fluorescence Spectroscopy鑑定 National Taiwan University Center for Nano Science and Technology http://nanost.ntu.edu.tw台灣大學奈米科技研究中心

  37. Colormetric Polynucleotide Detection Based on DNA-modified Gold Nanoparticles Probe Prior to addition of cDNA After Addition of cDNA Ref:C. MirkinJ. Am. Chem. Soc. 2003, 125, 1643-1654

  38. Selective Colorimetric Detection of DNA Based on the Distance-Dependent Optical Properties of Gold Nanoparticles Ref: C. Mirkin et al. Nature1996, 382, 607-609.

  39. No Pb2+ Pb2+ Color developed on an alumina TLC plate after 10 min of nanoparticle aggregation = 17E DNAzyme-Directed Assembly of Gold Nanoparticles and Their Application as Biosensors for Pb2+ Ref: Y. Lu et al J. Am. Chem. Soc. 2004, 126, 12298-12305

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