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Quantum dots as tools for cellular imaging

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Avidin or protein-G with positively charged tail ... In Xenopus. Dubertret et al. 2002. Quantum dots and imaging. In vivo visualization of capillaries ... – PowerPoint PPT presentation

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Title: Quantum dots as tools for cellular imaging


1
Quantum dots as tools for cellular imaging
  • Presented by
  • Debasish Sen
  • Department of Physiology and Biophysics
  • School of Medicine
  • University of California, Irvine

2
What are quantum dots ?
  • Crystalline fluorophores
  • CdSe semiconductor core
  • ZnS Shell

3
What are quantum dots
  • Unique Spectral properties
  • Broad absorption
  • Narrow emission
  • Wavelength depends on size
  • Hydrophobic crystals

4
Making hydrophobic quantum dots bio-compatible
  • Various methods for making them water-soluble
  • Derivatizing surface with mercaptoacetic acid
  • Encapsulating in phospholipid micelles
  • Derivatizing surface with DHLA
  • Coating them with amine-modified polyacrylic acid

5
Conjugating quantum dots to biomolecules
  • Avidin or protein-G with positively charged tail
    conjugated to negatively charged DHLA coat of
    quantum dots

6
Quantum dots v/s other fluorescent probes
  • Photostability (quantum dots do not photobleach)

Wu et al. 2003
Red qdot 605 Conjugate
Green Alexa488 Conjugate
7
Quantum dots v/s other fluorescent probes
  • Broader excitation spectrum and narrower gaussian
    emission spectrum
  • No spectral overlap between dots of different
    size better for multiplexing

Jaiswal Simon 2004
8
Quantum dots v/s other fluorescent probes
  • Brighter than other fluorophores

Quantum dots
Fluorescein
Larson et al. 2003
9
Quantum dots and imaging
In Xenopus
Dubertret et al. 2002
10
Quantum dots and imaging
Quantum dots
FITC-Dextran
In vivo visualization of capillaries
Larson et al. 2002
11
Quantum dots and imaging
Wu et al. 2003
12
Quantum dots and imaging
Diffusion of single Qdot-GlyRs in synaptic boutons
Glycine Receptors
Dahan et al. 2003
13
Quantum dots and imaging
EGF receptor
EGF-QD
Live imaging of receptor mediated endocytosis
Lidke et al. 2004
14
Quantum dots and imaging
Individual vesicles
Observing high resolution structure in dendritic
cells
15
Quantum dots and imaging
Individual vesicles -Temperature variations
Dendritic cell processes
Observing high resolution structure in dendritic
cells
16
Quantum dots and imaging
Single quantum dot crystals can be observed in
electron micrographs
17
Quantum dots and imaging
  • Quantum dots have been used in FRET
  • In conjunction with Texas Red
  • In conjunction with fluorescent quenchers

Willard et al. 2003
18
Advantages
  • Specific labeling of cells and tissues
  • Useful for long-term imaging
  • No cytotoxicity observed in vitro/in vivo
  • Useful for multi-color multiplexing
  • Suitable for dynamic imaging of subcellular
    structures
  • May be used for FRET-based analysis

19
Disadvantages
  • Colloidal polymer-coated quantum dots can
    aggregate irreversibly
  • Quantum dots are bulkier than many organic
    fluorophores
  • Accessibility issues
  • Mobility issues
  • Cannot diffuse through cell membrane
  • Use of invasive techniques may change physiology

20
Acknowledgements
Mark Ellisman Tom Deerinck NCMIR, UCSD
Mike Cahalan
Ian Parker
Lu Forrest Mark Miller Sindy Wei Melanie
Matheu Olga Safrina
UCBREP GREAT
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