W. Richard Bowen and Nidal Hilal 4

7.2 ENgINEERINg THE ECM FOR PRObINg CELL SENSINg 201
which react with the substrate. Upon SAM formation, the chemical properties
of the surface are no longer determined by the underlying substrate
but by the exposed functional tail groups at the non-substrate end of the
SAM. A commonly used system is the self-assembled monolayer of an
alkanethiol (SH(CH 2) nX) on gold; the sulfhydryl head groups (SH–) spontaneously
form sulphur–gold bonds and leave the functional tail group
structures (X) tethered away from the surface. The terminal X groups can
be tailored to have different functionalities [20], e.g., NH 2– and COOH–
groups for subsequent attachment of proteins or peptides via common
bioconjugation chemistries [21]. The use of mixed or diluted monolayers
and tuning the length of the hydrocarbon spacer chain can also be used to
achieve desired surface properties [22]. SAMs of alkylsilane on hydroxylated
surfaces of SiO 2/Si substrates, such as glass and silica, have also been
extensively developed, although here, greater care needs to be taken over
the reaction conditions to avoid self-polymerisation on the surface.
As photolithography requires clean room facilities to prevent dust particles
spoiling the pattern transfer process, the infrastructure costs mean that
these techniques are often not easily accessible on a regular basis. Thus, a
pioneering method has been developed by Whitesides’ group, called ‘soft
lithography’ (microcontact printing, Figure 7.2(B)). This uses a physical
stamp made from poly(dimethysiloxne) (PDMS) elastomer [23]. The stamp
is a negative replica of a microstructured substrate (master) made using
conventional photolithography and/or etching methods. Generally, the
stamp is fabricated by thermal curing of PDMS against a master template
followed by peeling away the cured structure. The PDMS stamps can then
be inked with silanes, alkanethiols or ECM proteins to produce patterns
on a wide range of substrates [24]. Unstamped regions can be blocked to
resist non-specific protein adsorption by various methods such as the use
of PEG-terminated SAMs or rinsing with denatured albumin solution. The
PDMS stamps can be repeatedly replicated from the master and reused
many times, thereby significantly reducing the cost of fabrication. Using
this methodology, microcontact printing has greatly extended the range of
substrates that can be engineered for use in cell studies since many of the
polymeric materials compatible with biological studies are not suitable for
patterning using traditional photolithography processes.
An early example of the use of micropatterned surfaces is one which
allowed systematic examination of the interaction of cells with substrate
surface chemistries and ECM components [25]. Observations of cell
growth on metallic and polymeric micropatterns have provided invaluable
information to develop new types of cell culture vessels and screen
the biocompatiblity of metal materials for implants. Cell function data on
SAM patterns have enabled the identification of chemistries and systems
that possess designed properties; e.g. observations from patterns made
using a range of poly(ethylene glycol) (PEG)-derivatised alkanethiols