Hydrogel-Based Microfluidic Cell Culture 97
5.2.6 N-isopropylacrylamide Polymers (NiPAAm)
N-isopropylacrylamide (NiPAAm) homopolymers (Fig. 5.6) and copolymers are
thermoreversible polymers which have been used for drug delivery and tissue
engineering. At low temperature, the polymer dissolves is a liquid solution since
hydrogen bonds form between polar groups in the polymer and water. The
polymer chains form extended coils surrounded by ordered water molecules. The
gel is observed at high temperature. There, the polymer precipitates out of solution
because hydrophobic interactions dominate, and the polymer chains transition to a
globular state.
36
NiPAAm homopolymer in pure water has a lower critical solution
temperature (LCST) of 32–34
C, where the transition occurs between liquid and
gel. NiPAAm copolymers which contain hydrophilic groups such as polyethylene
glycol (PEG) will increase the LCST. Reversible gelation occurs due to polymer
chain entanglements and the increased hydration.
In contrast to other thermally reversible systems such as Pluronics, NiPAAm
copolymers when gelled do not revert to the liquid state upon dilution. The
NiPAAm phase transition also occurs over a narrower temperature range, permit-
ting more precise control for on-chip gelation.
Copolymers of NiPAAm have already been used for 3-D culture of chondr-
ocytes
37
and pancreatic islets.
38
Acrylic acid or polyethylene glycol (PEG) copoly-
mers are used to increase the hydration of the gel, since NiPAAm homopolymer gel
exhibits high water loss. This change in hydration with temperature has been used
to fabricate cell sheets. When cells are cultured on surfaces treated with NiPAAm
at 37
C, the cells adhere to the slightly hydrophobic surface. When the temperature
is brought below the LCST, the gel swells and becomes more hydrated, and the
cells spontaneously detach. This effect has been used to create confluent sheets
of myocardial cells which can be layered to build 3-D electrically communicative
structures.
39
5.3 MICROFABRICATION
Microfabrication processes are the basis for integrated circuit (IC) manufacture.
They involve thin film deposition, patterning, and etching. In addition to fab-
rication of planar silicon-based ICs, microfabrication processes have enabled the
development of micro-electro-mechanical systems (MEMS), as well as an increas-
ing variety of glass- and polymer-based lab-on-a-chip and implantable devices for
biomedical applications.
More recently, the technique of soft lithography has been developed to cre-
ate microstructures using printing, molding, and embossing processes.
40
This
technique most commonly uses poly(dimethylsiloxane) (PDMS) as an elastomeric
mold material. Elastomeric PDMS structures are replicated from a mold master,
which is commonly either a silicon or glass substrate patterned with photolithog-
raphy (Fig. 5.7). If a high-aspect ratio photoresist such as SU-8 is used, the mold
master can be produced using only one photolithography step. Alternatively, the
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98 M. C. W. Chen and K. C. Cheung
D E
F G
H I
Figure 5.7. Microfabrication of PDMS structures. (a–b) Photoresist is spin-coated on a
silicon wafer. (c) A mask is placed in contact with the layer of photoresist. (d) The
photoresist is illuminated with ultraviolet (UV) light through the mask. An organic
solvent dissolves and removes photoresist that is not crosslinked. The master consists of
a silicon wafer with features of photoresist in bas-relief. An expanded view of one of the
microfabricated structures with its characteristic critical dimensions is shown. (e) PDMS is
poured on the master, cured thermally and peeled away. (f) The resulting layer of PDMS
has microstructures embossed in its surface.
47
For color reference, see page 261.
mold can be produced through a photolithography step followed by reactive ion
etching of the silicon substrate. PDMS is poured onto the mold master, cured,
and then removed. The PDMS structure can be used as a stamp for microcontact
printing of proteins or self-assembled monolayers. This PDMS structure can also
form closed microfluidic channels when bonded to another substrate.
PDMS has many advantages for use in biomedical applications: flexibility,
transparency, biocompatibility, and gas permeability. Prototype PDMS structures
can be made rapidly and at relatively low cost. Elastomeric structures made
in this method have been used in microstamping, microfluidic patterning, and
stencil patterning to create well-defined areas of cellular adhesives, polymers, self-
assembled monolayers (SAMs), and biomolecules to study cellular interactions on
the microscale.
41
PDMS microchambers have been used as stamps for patterning
cell attachment on surfaces,
42
chambers for DNA amplification via the polymerase
chain reaction (PCR),
4345
and as chambers for bacterial
46
(Fig. 5.7) and mam-
malian cell culture.
47
PDMS will swell and absorb liquid media, which can be a major consideration
for cell culture applications involving small volumes. The amount of liquid
absorbed depends on the ratio of siloxane oligomer precursors used to make the
PDMS.
48
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