Background:
It has been shown that uniform nanodiscs and nanovesicles can
self-assemble in phospholipid mixtures composed of long- and
short- chain lipids. Previously, we were able to manufacture
large quantities of such nanoparticles (NPs), which entrapped
the hydrophobic fluorescence dye (e.g., Nile Red), and stabilize
the NP structures by either charged or polyethylene glycol
conjugated (PEGylated) lipid. The original proposal is to apply
these NPs is for drug delivery carriers to target cancer cells. In
addition to this application, here we demonstrate another
potential application for biosensor using these NPs.
• The preparation protocols of uniform lipid-
based NPs are simple, robust, environmentally
friendly and scalable.
• The cellular uptake of nanodiscs is than that of
nanovesicles by CCRF-CEM , presumably due
to more internalization pathways are available
for discs than vesicles.
• The NPs can be used for instrument-free, high-
sensitivity and low-cost pathogen sensors.
Summary
Graduate Students:
Ming Li (IMS, UCONN), Andrew Hu (Mech.
Eng.), Yan Xia, Ying Liu (Chem. Biomol.
Eng.) and Wafa Aresh (Biomed. Eng.)
Scalable Manufacture of Multi-Functional Lipid-Based Nanoparticles
(Grant #: NSF-CMMI 1131587) Mu-Ping Nieh1, Tai-Hsi Fan1, Yong Wang2
University of Connecticut1, Pennsylvania State University2
Spontaneous Structural Diagram
Materials & Strategy
Zwitterionic long-chain lipids:
dimyristoyl (di-C14) phosphatidylcholine (DMPC) or
dipalmitoyl (di-C16) phosphatidylcholine (DPPC)
Zwitterionic short-chain lipid:
dihexanoyl (di-C6) phosphatidylcholine (DHPC)
PEGylated lipid:
Polyethylene glycerol (2000) - distearoyl (di-
C18) phosphatidylethanolamine (PEG-DSPE)
Charged long-chain lipids:
dimyristoyl (di-C14) phosphatidylglycerol (DMPG) or
dipalmitoyl (di-C16) phosphatidylglycerol (DPPG)
Fluorescence
Dye
(hydrophobic
molecules)
Increasing T
Nanodisc-to-Nanovesicle Transition
Decreasing T
Nanodiscs Spherical vesicles Oblate vesicles
Cellular Uptake CCRF –CEM (T-lymphoblastic leukemia)
Fluorescence Confocal Microscopy
• The cell uptake of nanodiscs is 3 ~ 5 times of that of nanovesicles.
Fluorescence 0 mm
0 mm
3 mm
3 mm
5 mm
5 mm
7 mm
7 mm
Fluorescence Flow Cytometry
Cel
l co
un
ts
Fluorescence Int.
Cellular
Uptake
Mechanism
Inhibitors Inhibited Mechanism
Chlorampramazine Clathrin
Filipin III Caveolae
Cytochalasin D Macropinocytosis
Amiloride Macropinocytosis
Wortamannin Macropinocytosis
Nocodazole Microtubules
0
20
40
60
80
100
120
Med
ian
flo
ure
cen
ce u
pta
ke
(% o
f co
ntr
ol)
Discs Vesicles
Clathrin
Caveolae
Macrpinocytosis Microtubules
Vesicles only take Clathrin- and
Caveolae-mediated pathways, while
nanodiscs take all four pathways .
(1) Drug Delivery Carrier
(2) Biosensing
Induced NP Aggregation by
polymer linker
Individual nanodiscs
50 nm
“strung” nanodiscs
100 nm
“strung” vesicles
Addition of the Polymer Linkers
high charge
density
low charge
density
108 CFU/mL: 104 102 0
Instrument-free Pathogen
Sensing
No
coverage }
