Lab on a chip used for protein studies
Ten years ago, ORNL's Mike Ramsey built the first lab on a chip. Now, improved
versions of this miniature chemistry lab are being shipped all over the world. A
toaster-sized, computerized device containing four matchbox-sized protein identification
chips modeled after Ramsey's invention is being mass-produced by Caliper
Technologies, Inc., in California. It is being sold to biotechnology firms by Agilent
"Proteins, like DNA, pose a massive chemical measurement problem," Ramsey says.
"But we have learned how to use a lab on a chip to measure molecular weights of
proteins in much smaller samples and in much shorter times than are required by
Caliper Technologies is developing microchips for drug discovery. Such a device would
help pharmaceutical firms rapidly identify compounds effective in inhibiting the activity
of disease-causing proteins. "We believe that an automated device with massively
parallel microfluidic chips can work with sample volumes that are 1/10,000th the
volumes analyzed in conventional benchtop devices, at 10 to 100 times the speed or
more," Ramsey says.
The current drug discovery chip contains four channels—thinner than human hair—that
connect reservoirs, all of which are carved into a rectangular glass plate, using
microfabrication technologies. A disease-related enzyme is introduced into a chip
channel. Because of pressure differences, the enzyme and a modified substrate flow
through the channel network, mix, and react. The reaction product is fluorescent when
exposed to a laser beam. The amount of fluorescence is a measure of the reaction rate.
When a test compound is introduced into the chip through another channel, it typically
reacts with the enzyme, blocking out the substrate so less of the fluorescent product is
produced at a time. The reduced fluorescence signal indicates the effectiveness of the
test inhibitor compound. By introducing different test compounds to the device every 5
seconds, it is possible to rapidly compare reaction rates to identify potentially effective
drugs. In a recent demonstration at Caliper, nearly a million compounds were screened
while using less than 1 microgram of enzyme (usually a very valuable material).
In ORNL's Chemical and Analytical Sciences Division, considerable research is being
conducted by Ramsey's group on developing improved lab-on-a-chip technologies for
biological, environmental, forensic, and defense applications. The lab on a chip has
been honored by R&D magazine as one of the 40 top innovations that have come about
since the magazine began its R&D 100 competition in 1963. It also has been recognized
by a panel of citizens as one of the top 23 technologies developed using Department of
In 1998 at Caliper Technologies, Rose Ramsey (Mike's wife) and a colleague there first
demonstrated that the lab on a chip could complete a two-dimensional (2D) separation
of peptides in under 10 minutes. The 2D chip separation uses two types of separations
to resolve the peptides. In one channel, they are separated by differences in migration
speed and in another channel by differences in peptide charge and size in response to an
electric field (capillary electrophoresis). By contrast, it takes 24 to 48 hours to do this
separation using conventional 2D gel electrophoresis. More recently, Stephen Jacobson,
Chris Culbertson, and Norbert Gottschlich contributed to a newly designed 2D chip that
works even faster.
Mike, Rose, and Robert Foote
of ORNL then received funding
from the Laboratory Directed
Research and Development
Program at ORNL to analyze
proteins by combining the lab
on a chip with an electrospray
ionization time-of-flight mass
spectrometer (ESI/MS). Rose,
who conceived the idea, and
Iulia Lazar, a post-doctoral
fellow at ORNL, showed that
the procedure could be used to
rapidly analyze hemoglobin, the
protein that makes blood cells
red and transports oxygen from
the lungs to the body tissues.
In the chip, the hemoglobin
from a drop of blood is reacted
with the enzyme trypsin, which cleaves the blood protein at the scattered sites of two
amino acids. The fragments of the hemoglobin are electrosprayed directly from the chip
as ions into the mass spectrometer.
"The pattern of the fragments gives a fingerprint for the protein that can be compared to
amino-acid sequences in the protein database, allowing us to identify the protein," Rose
says. She used this technique to sequence over 70% of human hemoglobin from a drop
of blood. She demonstrated that the technique can rapidly distinguish sickle-cell
hemoglobin, whose fingerprint is different from that of normal hemoglobin. This work
was recently published in Analytical Chemistry. This technique could also be used to
rapidly screen for other hemoglobin variants.
The lab on a chip may be small but its potential for advancing medical diagnosis and
treatment is quite large.