FUNCTIONALIZED COLLOIDAL NANOPARTICLES

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FUNCTIONALIZED COLLOIDAL NANOPARTICLES

• Jaan Noolandi

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BACKGROUND

• Proteome PROTEins expressed by a genOME
• Study behavior of system to produce a living
  organism
• Complexity requires new analytical tools for
  greater
• versatility and higher throughput than
  existing methods
• Emphasis on multiplexing and miniaturization
• Current Methods
• Protein microarrays (chips)
• Self-encoded latex beads
• Submicrometer metallic barcodes
• Functionalized nanoparticles

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General flow scheme for proteomic analysis
separation

• Protein
• mixture

Proteins
digestion
digestion
Peptides
MS analysis
Protein mixture
separation
Database search algorithms
MS data
identification
Introduction to Proteomics - Tools for the New
Biology Daniel C. Liebler, Humana Press (2002)
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• Mass Spectrometry and
• Proteomics
• Steve P. Gygi and Ruedi
• Aebersold
• Current Opinion in
• Chemical Biology 4 ,
• 489-494 (2000)

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• Protein Arrays
• New Focus, Science 289, 1673 (2000)

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Optical Coding of BiomoleculesShuming Nie et
alNature Biotechnology 19, 631-635 (2001)
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Synthesis of barcoded particles.
Submicrometer Metallic BarcodesChristine D.
Keating, Michael J. Natan et alScience 294,
137-141 (2001)
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• Synthesis of Functional Colloidal Nanoparticles
• Monodisperse arm lengths to obtain well-defined
  mobility of
• bare particle
• Optimize arm lengths
• Optimize arm density

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Capillary Electrophoresis
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Bioaffinity Capillary Electrochromatography
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protein extract
Probe-proteincomplex
separateby capillary electrophoresis
Functionalized colloidal particle probe
Digest,MS analysis
Identify
Bait approach to detection of DNA-interacting
protein with a functionalized colloidal particle
probe.
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SYNTHESIS STRATEGY

• Hydrophobic / hydrophilic block copolymer
• - hydrophilic block (sulfonated polystyrene,
  maleic anhydride)
• - hydrophobic block (polystyrene)
• Micelle formation
• - add monomer, initiator
• - form miniemulsion
• - polymerize
• Photo-emulsion polymerization
• - core latex particles by emulsion
  polymerization
• - add polymerizable photoinitiator
• - polymer brush by emulsion polymerization

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Conjugation of Biomolecules with Synthetic
Polymers
O
O
N
SH
NH(CH2-CH2O)nH
FAM-(dT)20 or 35
O
O
O
S
N
FAM-(dT)20 or 35
NH(CH2-CH2O)nH
O
Reaction of a maleimide-terminated PEG polymer
with a 5-thiol-terminated ssDNA oligomer

• end functionalize water-soluble polymer with
  maleimide group

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SUMMARY

• Increase throughput of proteomics assays by
  combining
• capillary electrophoresis involving colloidal
  particles
• characterized by well-defined molar mass
  profile with
• massive parallel processing by capillary
  arrays
• Colloidal particles synthesized by different
  synthetic
• routes (photo-emulsion polymerization,
  micelle
• formation, living radical polymerization)