Biomedical Engineering Reference
In-Depth Information
Amy Herr's laboratory at the University of California (Berkeley) has developed a microlu-
idic glass chip platform that integrates polyacrylamide gel electrophoresis with immunoblot-
ting (
Figure 4.25
). he main chamber is illed with gels that are photochemically patterned to
provide either separation or immunoblotting functionality (
Figure 4.25a
). Transport of luids
within the chip is achieved by electrophoresis; to achieve homogeneous current injection and
parallel electric ield lines, the main chamber is designed with many electrical contacts (con-
nected to ports numbered “4” through “8” in
Figure 4.25b
). he sample (5 μL) is loaded onto
reservoir 2 and moved sequentially down and right by applying voltage (50-500V) sequentially
to the various ports (
Figure 4.25c
). he yellow band stops at (i.e., is captured by) the antibody-
functionalized blotting region despite the continued application of ield: note that the blue band
(not captured by the antibodies) keeps going out of the ield of vision. An assay takes typically
1 minute and the chips can be reused ater cleaning with piranha solution.
4.6.7 Protein Crystallization Chips
Elucidating the relationships between protein structure and protein function has been one of the
most intense areas of study in the ield of molecular biology for many decades, and it remains so
in this decade of high-throughput experimentation where it has become one of the goals of pro-
teomics. he determination of a protein's structure is presently done by X-ray crystallography
and requires the formation of a crystal of the protein so that an X-ray spectrum (the set of X-ray
dots scattered by the crystal onto a screen) can be recorded and analyzed. Protein crystal forma-
tion depends on a number of biophysical parameters that are typically optimized empirically,
for example, temperature, salt and protein concentration, and others. Not all conditions produce
good crystals. Screening this combinatorial parameter space may involve running on the order
of hundreds of experiments, which has slowed the progress of several ields that rely heavily on
protein structure determination, such as structural biology, molecular biology, and pharmacol-
ogy. In 2002, Stephen Quake's group (then at Caltech) produced a chip with 480 microvalves
capable of simultaneously performing 144 mixing reactions (each using only 10 nL of protein
sample). Ater incubation at 25°C (for up to 3 weeks), the resulting crystals (
Figure 4.26
) could
be removed from the chip and analyzed. he microluidic chip is now commercially available.
a
b
c
100 µm
d
e
f
FIGURE 4.26
Protein.crystallization.on.a.chip..(a-e;.From.Carl.L..Hansen,.Emmanuel.Skordalakes,.
James. M.. Berger,. and. Stephen. R.. Quake,. “A. robust. and. scalable. microluidic. metering. method.
that.allows.protein.crystal.growth.by.free.interface.diffusion,”.
Proc. Natl. Acad. Sci. U. S. A.
.99,.
16531-16536,. 2002;. f,. from. Carl. L.. Hansen,. Morten. O.. A.. Sommer,. and. Stephen. R.. Quake,.
“Systematic. investigation. of. protein. phase. behavior. with. a. microluidic. formulator,”.
Proc. Natl.
Acad. Sci. U. S. A.
.101,.14431-14436,.2004..Copyright.(2002.and.2004).National.Academy.of.
Sciences,.U..S..A..Figure.contributed.by.Stephen.Quake.)
