Transcript Document

Magnetic silica spheres
with large nanopores
for nucleic acid
adsorption
and cellular uptake
Jian Liu, Bo Wang, Sandy Budi Hartono
Biomaterials
University of Queensland, Australia
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contents
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Introduction
Experimental Section
Results and Discussion
Conclusions
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Introduction
Mesoporous materials
 Large specific surface area
 Large pore volume
 Uniform pore size distribution
 Catalysis
 Imaging
 Drug delivery
Mesoporous silica nanoparticles (MSNs)
 Biocompatibility
 Low toxicity
 Biological application
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Introduction
Other properties that MSNs required for
biological application
 Large pore sizes
 Appropriate magnetic properties
 Appropriate functional surface
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Introduction
1、Large pore size
 Large internal surface
 Large mesoporous volume
 Cytochrom C
2.6×3.2×3.3 nm
 a-L-arabinofuranosidase
3.9×9.7×14.4 nm
Suitable pore sizes for immobilisation of these proteins
can vary from10 to 50 nm
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Introduction
2、Magnetic properties
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Bioseparation
Cell sorting
Diagnostic analysis
Simultaneous imaging and drug delivery
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Introduction
Preparation methods：
 Large pore size mesoporous materials
Templates：Pluronic P123
Swelling agent：1,3,5-trimethyl benzene (TMB) or alkanes
Condition：Strong acidic
 Magnetic mesoporous materials
Templates：Brij56 micelles
Condition：Basic
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Introduction
3、Functional surface
To delivery nucleic acids, the silica surface
with positive charge is needed to electrostatically
bind DNA and RNA molecules
Methods：
 Functionalisation with amine-derivative
group such as APTES
 Conjugations with cationic polymers such as PEI
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Introduction
 Develop synthesis methods to prepare MSNLP
 Establish a surface functionalisation method to
enable adsorption and delivery of nucleic acids
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Experimental Section
Synthesis of monodisperse superparamagnetic Fe3O4
nanocrystals
1-octadecene
1,2-hexadecanediol
+
Static conditions at 250℃
in a Teflon-lined autoclave
for 6~12 h
The concentration of the magnetic
nanocrystals is 10 or 30 mg/ mL
and suspended in hexane
Fe3+
Iron stearic acid
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Experimental Section
Synthesis of magnetic silica nanospheres with large
nanopores
PLL
30-glycidox-ypropyltrimethoxysilane (GOPS)
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Experimental Section
DNA adsorption
 CpG DNA 1826 (5‘ to 3‘, TCCATGACGTTCCTGACGTT）
 Measuring A260 absorbance at 260 nm
Transfection of cells
 CyTM3-labeled miRNA
 Rat kidney epithelial cells (NRK-52E)
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Results and Discussion
Synthesis of magnetic silica nanospheres with large
nanopores
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Results and Discussion
Fig. 1. SEM (a, c), TEM (b, d-f), and HRTEM (g, h) images of MSNLP synthesised with different amount of
hexane: MSNLP-0-350 (a, b), MSNLP-10-350 (c, d), MSNLP-10-700 (e), MSNLP-10-1400 (f-h).
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Results and Discussion
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Results and Discussion
N0I0 route
 Brij56: polyoxyethylene 10 cetyl ether, C16H33EO10
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Results and Discussion
Magnetic properties of magnetic silica nanospheres with
large nanopores
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Results and Discussion
Fig. (A) Field-dependent magnetisation at 300 K of MSNLP with different amounts of magnetite: a)
MSNLP-10-350, b) MSNLP-10-700, c) MSNLP-10-1400, and d) MSNLP-30-1400; and (B) the separation
process of MSNLP-30-1400 nanospheres from solution by magnet (right picture) and their re-dispersion
by as slight shake (left picture).
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Results and Discussion
Composition of PLL functionalised MSNLP
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Results and Discussion
Adsorption of DNA on PLL functionalised
magnetic silica nanospheres with largenanopores
MSNLP-0-350-PLL qm=22.5μg/mg
MSNLP-10-350-PLL qm =15μg/mg
MSNLP-10-1400-PLL qm=10μg/mg
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Results and Discussion
Fig. Left panel a-d, cells
transfected with fluorescent
oligonucleotide only; middle panel
e-h, cells transfected with
nanoparticles alone; and
right panel i-l, cells transfected with
nanoparticles loaded with
fluorescent oligonucleotide. From
top to bottom: cy5 channel images of fluorescence of CyTM3
labeled miRNA (red),
F-actin - images of F-actin stained
by FITC-Phalloidin (green), DAPI images of nuclei stained with DAPI
(blue), and merge - the merged
picture.
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Conclusions
 Magnetic silica nanospheres with large nanopores(13-20
nm) were synthesised for the first time
 The saturation magnetisation values can be conveniently
controlled by changing the amount of Fe3O4 magnetic
nanocrystals encapsulated
 After functionalisation with PLL, high adsorption capacity
ranging from 10 to22.5 μg/mg for CpG DNA and efficient
cellular delivery capability for miRNA were achieved
 The materials synthesised in this study could find broad
applications
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Thank You
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