MOON'S RESEARCH HOME (under construction)
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Dr. Moon received his Ph.D. in inorganic
and materials chemistry from Pohang University of Science and Technology
(Korea) in August 1999, where he studied surface modifications and
characterizations.
He optimized aminosilane layers for efficient
target oligonucleotide hybridization in microarray experiments.
He also established a facile and straightforward
quantification method for amine density.
At MIT, Dr. Moon studied the synthesis of new poly(phenylene ethynylene) polymer brushes and the effect of unique surface structures on chemical sensing under Prof. Tim Swager. In 2001, Dr. Moon joined the Nomadics Life Sciences/Advance Materials Laboratory in Cambridge, MA. Dr. Moon established various fabrication and modification methods of amplifying fluorescent polymers (AFPs) for efficient sensory platforms (film, sol-gel, and hybrid materials). He was a visiting scientist at the Institute for Soldier Nanotechnology at MIT. After seven-year industrial experiences, Dr. Moon moved to the Department of Chemistry at Florida International University. His current research focuses on the development of novel fluorescent materials for sensitive and reliable detection of target molecules. Conjugated polymer nanoparticles are currently developing for labeling of cancer cells at tissue levels and delivery of therapeutic agents to targeted cells. He is also interested in developing nanostructured materials and surfaces for sensitive detection of trace chemicals such as explosives or drugs. |
1. Conjugated Polymer Nanoparticles for Fluorescent Imaging
Fluorescent labeling and detection of target biological molecules in live cells is an essential way of studying complex and dynamic cellular processes. Many fluorescent dyes and engineered fluorescent proteins are widely used for these applications due to their small size and biocompatibility. However, poor photostability of these probes limits their broad applicability in long-term monitoring of live cells with high sensitivity. Quantum dots (QDs) are considered as an alternative probe owing to their excellent optical properties such as high photostability, narrow emission, and high brightness. However, the inherent toxicity of QDs (mainly from a heavy metal core such as divalent cadmium ions) causes concern in long-term monitoring of cellular events. In addition, difficulties associated with surface modification of QDs also retard their applications in live cell systems. Therefore, novel materials that overcome the stability and toxicity issues in live cell imaging are in high demand.
Conjugated polymers (CPs) are attractive fluorescent materials that address the requirements for fluorescence microscopic imaging. CPs are organic materials that are readily synthesized by well established synthetic chemistries using metal catalytic systems. The fluorescent nature of CPs originates from the conjugated π-electrons, which travel efficiently along the aromatic backbone. Characteristic photophysical features of CPs are high fluorescence quantum yields, large extinction coefficients, efficient and fast energy transfers, and nonlinear optical behaviors due to the large two-photon absorption (TPA) cross-section.
Despite the promising characteristics, optical imaging applications of CPs in cellular, tissue, and animal systemsare largely limited by the intrinsic hydrophobic nature. Aggregation between polymer chains and nonspecific adsorption of proteins are the most obvious limitations for imaging in biological systems. Due to the limited aqueous solubility of the charged CPs, they tend to form aggregates in water or highly saline solution. The limited solubility is also a hurdle for the post modification of CPs for coupling of proper sensing elements or targeting agents.
The
challenges are to preserve the attractive photophysical properties of CPs in
aqueous phases and manipulate the physical properties of CPs so that they become
more suitable for biological applications. We have demonstrated that organic acid treatment of CPs followed by ultrafiltration produces water-dispersed conjugated polymer nanoparticles (CPNs) exhibiting high quantum yields, photostability, and non-toxicity to live cells.
The key discovery is that the controlled chain-chain interaction by organic acid allows stable nanoparticle formation while the photophysical properties of CPs are preserved in aqueous phases. Further fine tuning of the particle formation leads to dramatic reduction on the particle size below 10 nm.
Live
cell imaging application: An amine-containing poly(phenylene ethynylene)
(PPE) was designed and fabricated into CPNs, and the resulting CPNs were used
for live cell imaging. Various cells, including BHK (babyhamster kidney)
and BALB/C 3T3 (mouse embryonic fibroblast), were incubated with the CPNs in
culture media for various time periods(from 1 h to several days) to examine the
cellular uptake, photostability, and cellular toxicity of the CPNs. Figure shows
microscopic images of live (a) and fixed (b)
BALB/C 3T3 cells, stained by the CPNs overnight.
The CPNs accumulated randomly throughout the
cytosol. Confocal microscopic studies of fixed cells further suggests that CPNs
accumulated in some vesicular structures such as early or late endosomes.
The CPNs are up-taken by live cells without any measurable inhibition of cell
viability. In addition, CPNs exhibit high resistance to photobleaching, in
contrast to commercially available dyes.Current our research focuses on 1) the physical and photophysical optimization of CPNs by rational material designs and fabrications; and 2) the bioconjugation of CPNs with targeting agents for the target specific labeling.
2. Extremely Bright Multiphoton Imaging Probes
Fluorescent molecules can be excited by absorbing photons of certain energy(e.g. 400 nm) and the excited molecules release the absorbed energy as fluorescence emission. If the excitation light source carries extremely high photon density, the molecules can also be excited by simultaneous (within the interval of 10XE-18 sec) absorption of two-photons (2P) of lower energy (i.e. 800 nm). Because of the requirements for the 2P excitation, pulsed femtosecond lasers are often used to focus the light to generate significant density of excitation lights. This allows selective excitations of the fluorophores at the focal point that supports various applications including deep tissue microscopic imaging.
2P imaging holds great promise for in vivo microscopic physiological studies in areas such as neurobiology, immunology, and tissue engineering. Advances in 2P endomicroscope design have further demonstrated the possibility of developing non-invasive diagnostic procedures for the detection of malignancy in organs such as the oral cavity and the intestine. Many of these applications are currently limited by a lack of fluorescent probes with large 2P action cross-sections. The availability of improved 2P fluorescent probes will enable tissue imaging with higher sensitivity, greater penetration depth, reduced tissue photodamage and tissue autofluorescence backgrounds, and lower instrument cost. Despite a number of previous efforts to design and synthesize new organic fluorophores with high 2P action cross-sections, the utility of these probes in biomedicine has been limited by their hydrophobicity and cytotoxicity. A recent development is the realization that quantum dots (QDs) have large 2P action cross-sections on the order of 10,000 Goeppert-Mayer units (GM). These QDs exhibit high photostability and narrow emission spectral width. Despite their excellent photophysical properties, broad biomedical applications of QDs are limited by drawbacks such as the existence of “dark” dots, heavy metal core-related cytotoxicity, and difficulties in surface modification. Therefore, the synthesis and fabrication of alternative probes with high 2P action cross-sections is a priority.
Our initial experiments indicate that CPNs exhibit extremely large 2P action cross sections, comparable photostability to QDs, and non-toxicity.
The
measured 2P action cross-section of the CPNs as a function of excitation
wavelength is shown in Figure. Our measurements show a cross-section that
ranges between 1,000 and 11,000 GM with a maximum at about 730 nm. These
cross-sections are at least 2–3 orders of magnitude higher than conventional
organic fluorophores. In addition, at wavelengths below 815 nm, CPNs have higher
cross-section values than some QDs. It is worth clarifying that the
cross-section value of CPN is per particle and is not normalized in terms of
number of repeating units in the PPE used to fabricate the CPNs. This high
cross-section ensures that CPNs can be imaged in cells without simultaneous
excitation of NAD(P)H, an endogenous co-enzyme associated with oxidative
photodamage in biological specimens. Although CPNs are excited at a
spectral range similar to NAD(P)H, CPNs have cross-sections approximately five
orders of magnitude larger than those of native fluorescent coenzymes. These
high cross-section values support the utility of CPNs as 2P probes, especially
considering the invariability of CPN cross-sections over extended storage
periods of at least three months at room temperature.
To examine the
photostability of CPN in biological systems, we incubated live BALB/C 3T3
fibroblasts with CPN and QD525 (Qtracker 525, Invitrogen), respectively.
Cells were fixed with paraformaldehyde,
and samples were continuously imaged at 780 nm with a power of 2 mW for a
one-hour period. The total dosage at
each pixel was 0.1 mJ. After background
rejection, the photobleaching decay curves (Figure)
showed that the CPNs and QD525 bleaching rates were virtually identical; CPN and
QD525 retain 86% and 83% of their initial intensities, respectively. Focusing on
only the 25 most intense pixels in the images, the photobleaching in these high
intensity regions is more severe in both cases with CPN and QD525 retaining 70%
and 60% of the initial intensity, respectively.
It is interesting to note that the brighter pixels bleach faster, which
is consistent with the presence of a higher local density of reactive oxygen
species. Similar photostability was
observed in CPN and QD during the hour of continuous irradiation, indicating
high resistance to bleaching in CPN that is comparable to QDs.
Human adult dermal
microvascular endothelial cells (HMVEC-d) were pre-incubated with CPNs followed
by CellTracker Red before seeding into the device.
PBS-washed cells were seeded and cultured in the device over a period of
four days. Within 24 h after cell
seeding, a concentration gradient of vascular endothelial growth factor (VEGF)
and sphingosine was set up across the collagen gel region in order to promote
capillary formation. The
2P excitation images (figure) show projection of capillary structures through 80
mm
of the central gel region (the z-stack consists of 40 images at 2
mm
step). Cells invade the collagen gel
region over three days and form initial sprouts that evolve into capillaries
with lumens.
Over the three imaging days (and over the five days since
initially added to the cells), both the CellTracker Red dye and CPN probes
persist in the cells, allowing continuous tracking and monitoring.
Addition of CPN probes has not stunted
the growth of these capillaries, proving that cell behavior is unaffected by CPN
addition, as has also been shown in previous cell viability studies.
The observed non-toxicity of CPN and its
longevity inside cells demonstrate its suitability as a long-term intracellular
marker. These results indicate that CPNs
could be useful for a broad range of applications including understanding immune
cell trafficking in animal models and monitoring implanted stem cell migration.
Current research activities include 1) further optimization of 2P characteristics by controlling polymer chain-chain interactions; and 2) target specific labeling of cancer cells at tissue level (collaboration with Prof. Peter So at MIT).
3. Chemical/Biological Sensors
4. Novel Delivery Vehicles for Therapeutic Agents
PUBLICATIONS (SELECTED, 2000-)
- A. Abdul Rahim, W. McDaniel, K. Bardon, S. Srinivasan, V. Vickerman, P. T. C. So, and J. H. Moon, “Conjugated Polymer Nanoparticles for Two-photon Imaging of Endothelial Cells in a Tissue Model”, Adv. Mater. In print (2009)
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- J. H. Moon, Paul MacLean, William McDaniel and L. F. Hancock, “Conjugated polymer nanoparticles for biochemical protein kinase assay”, Chem. Commun.,
- J. H. Moon, William McDaniel, Paul MacLean and L. F. Hancock, “Live cell permeable poly(p-phenylene ethynylene)”, Angew. Chem. Int. Ed., 46, 8223 (2007)
- J. H. Moon, William McDaniel, and L. F. Hancock, “Facile fabrication of poly(p-phenylene ethynylene)/colloidal silica composite for nucleic acid detection”, J. Colloid Interface and Science, 300, 117 (2006)
- J. H. Moon, R. Deans, E. Krueger, and L. F. Hancock, “Capture and Detection of a Quencher Labeled Oligonucleotide by Poly(phenylene ethynylene) Particles”, Chem. Commun. 104 (2003)
- J. H. Moon and T. M. Swager, “Poly(p-phenylene ethynylene) Brushes”, Macromolecules, 35, 6086 (2002)
- T. –H. Kang, K. –J. Kim, C. –C. Hwang, K. Ihm, H. –J. Shin, M. –K. Lee, B. Kim, Y. –H. La, J. H. Moon, H. J. Kim, J. W. Park, C. –Y. Park, “Selective Cleavage of Functional Groups in Functionalized Organic Monolayers by Synchrotron Soft X-ray”, Surface Review and Letters,
- H. J. Kim, J. H. Moon and J. W. Park, “A Hyperbranched Poly(ethyleneimine) Grown at the Surfaces”, J. Colloid and Interface Sci., 227, 247 (2000)
- J. H. Moon, Y. –H. La, J. Y. Shim, B. J. Hong, K. –J. Kim, T. –H. Kang, B. Kim, H. Kang, and J. W. Park, “Selective Cleavage of the Carbon-Halide Bond in Substituted Benzaldimine Monolayers by Synchrotron Soft X-ray: Anomalously Large Cleavage Rate of the Carbon-Bromide Bond”, Langmuir, 16, 2981 (2000)
PATENTS(SELECTED)
-J. H. Moon, “Species for clinical imaging”, US Patent 7,521,232 (2009)
-J. W. Park, Y. H. La, J. H. Moon, B. Kim, T. H. Kang, K. J. Kim, “Method for high resolution patterning using soft X-ray, process for preparing nano device using the same method”, US Patent 7,267,932 (2007)
-R. Deans, L. F. Hancock, J. H. Moon, J. R. Malayer, C. R. Clarke, J. M. Clarke, A. Ramachandran, “Luminescent polymers and methods of use thereof”, US Patent Application 20060024707 (2006)
-J. H. Moon and J. W. Park, “Method for determining density of amine groups in aminosilane layer”, Japan Patent 3706687 (2005)
TEACHING
-Special topics in organic chemistry (2008 Fall)
-Organic Chemistry I (2009 Spring)
-Spectroscopic Techniques (2009 Fall)