|Salivary Diagnostic Device Shows Promise
Researchers supported by the National Institute of Dental and
Craniofacial Research (NIDCR), part of the National Institutes
of Health, have engineered a portable, phone-sized test that in
minutes measures proteins in saliva that may indicate a developing
disease in the mouth or possibly elsewhere in the body.
The point-of-care test, one of several saliva-based diagnostic
devices now under development with NIDCR support, one day in the
future could become a common sight in the dentist’s office. As
envisioned by the researchers, a dentist would collect a small
saliva sample with a patient’s consent, load it into the diagnostic
cartridge, start the assay, and have a read out waiting after a
cleaning or a dental procedure has been completed.
Called IMPOD, the device is described in the March 27 issue of
the Proceedings of the National Academy of Sciences. In
the report, the scientists offer the results of proof of principle
experiments in which IMPOD reliably measured the concentrations
of MMP-8, an enzyme associated with chronic inflammation of the
gums called periodontitis.
“The IMPOD is designed to measure up to 20 analytes, or biochemicals,
at once,” said Dr. Anup Singh, a chemical engineer at the Sandia
National Laboratories in Livermore, Cal. and senior author on the
paper. “We haven’t scaled up to that point, but we are doing multi-analyte
analyses in the laboratory. Our greatest need right now is validated
biomarkers to enable further clinical studies. The basic engineering
of the device has been completed.”
According to Singh, he and his colleagues were intrigued a few
years ago by the many potential advantages of saliva as a diagnostic
fluid. These include easy collection, no painful needle sticks,
portability of the tests, and potentially a lower cost to patients
than blood assays.
But they were initially daunted by the research task at hand. “Saliva
is a mirror of blood, but with a caveat, said Singh. “It’s not
an exact mirror, meaning everything that is present in blood is
present in saliva but at concentrations 1,000 to 10,000 times lower.
It’s diluted by saliva and the other secretions in the mouth. So
we needed sensitivity 1,000 to 10,000 times better than we’d need
if we were screening serum samples.” Sensitivity refers to the
lowest amount of an analyte that a test can detect.
Singh said he and his colleagues chose to use a lab-on-a-chip
device. The term refers in this case to a microchip, roughly the
size of a laboratory slide, containing networks of tiny channels
and chambers in which the salivary assay, or biochemical test,
is performed. For the researchers, the challenge was to design
the entire assay to fit into the microchip. The scientists also
had to miniaturize the components required to run the microchip,
such as the power supply and optical detection unit, and integrate
them into a rapid, fully automated diagnostic system.
The result is the IMPOD, short for Integrated Microfluidic Platform
for Oral Diagnostics. The process begins with a series of microwells,
each as distinct as fingers on a hand. One well holds the saliva
sample, while the other wells contain cleansing buffering solution
and antibodies that are preprogrammed to bind the specific protein
of interest in saliva. The antibodies are tagged with a fluorescent
dye that can be illuminated and measured at the end of the assay.
With the punch of a button, the contents of the wells are released
and merge into a single channel about 40 microns wide, or roughly
the width of a human hair. As the mixture flows in these tight
quarters, the antibodies readily find the proteins of interest,
tag them, and continue forward to be trapped on a porous gel membrane
that serves as a filter.
“The bound proteins stack against the membrane because they are
too big to squeeze through the pores,” said Dr. Amy Herr, also
at Sandia National Laboratories and lead author on the paper. “Conversely,
the many smaller molecules in saliva flow right through the pores
and are filtered out and routed to a waste chamber.”
Thereafter, a voltage reversal releases the trapped proteins from
the gel. They continue down a channel, where a standard diode laser
shines onto the fluorescent tags and quantifies the concentration
of the protein in the sample. The dentist reads the result and
determines whether the protein levels correlate with a given disease.
To put their lab on a chip to the test, they collected saliva
from 23 people — 14 with periodontitis and nine in good oral
health. Loading roughly one-tenth of a drop of saliva for analysis,
the IMPOD processed the samples and produced a result in less than
five minutes. The results showed that on average those in good
oral health had lower concentrations of MMP, while people with
periodontitis on average had elevated levels of the tissue-damaging
enzyme. The results were confirmed with a standard ELISA blood
test, currently the gold standard.
“The data correlate nicely with those from other types of studies
that show that MMP-8 is indicative of periodontitis,” said Dr.
William Giannobile, a researcher at the University of Michigan
School of Dentistry in Ann Arbor. “There is every reason to believe
additional biomarkers for periodontitis can be identified, as the
work to catalogue salivary genes and proteins nears completion
and our understanding of periodontal disease continues to advance.”
The researchers noted that IMPOD, although developed for saliva,
could be used to test other diagnostic fluids such as blood and
The National Institute of Dental and Craniofacial Research is
the Nation’s leading funder of research on oral, dental, and craniofacial
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