- Scientists find new approach to optically trapping and manipulating small particles
Laser creates electrical field that can move particles
Optical trapping supports a variety of applications throughout the
biomedical research arena. Using light beams to create force gradients, it enables
investigators to trap and control suspended particles and even to build three-dimensional
nanostructures. But the technique is limited in application: It cannot be used alone
with particles smaller than the wavelength of light, including DNA fragments, oligonucleotides,
proteins and peptides.
Researchers have developed methods to overcome
this limitation. With one, for example, they attach the smaller molecules to larger
particles called “handles.” Still, the technique is hampered by the
practical issues of controlling larger particles.
Investigators with Protein Discovery
Inc. of Knoxville, Tenn., California Institute of Technology in Pasadena and Oak
Ridge National Laboratory in Tennessee wanted to find a way to move proteins
without using larger particles.
“Separating proteins according
to their type is very important in identifying proteins in a mixture,” said
Thomas Thundat of the national lab. “For example, a sample of serum will have
hundreds of different types of proteins. And transporting proteins is a first step
in separating them.”
Thundat and his colleagues at Oak Ridge
— including Robert J. Warmack and Gilbert M. Brown — therefore developed
a technique they call photoelectrophoretic localization and transport. It uses beams
of light incident on a photoconductive semiconductor surface to create controlled
electric field traps, allowing them to transport charged molecules across the surface.
Thundat likens this to moving the proteins through microfluidic channels, with an
“In our case, we do not have
any prefabricated channels,” he said. “The path of light on the surface
is the channel. That means we can move the molecules in any arbitrary path we like.”
The technique requires a photoconductive
electrode and a counterelectrode, which can be a small-area electrode, a large conductive
surface or even another photoconductive surface. They are kept in contact with an
electrolyte containing the molecules to be moved. An external circuit applies voltage
between the electrode and countelectrode, and this in turn creates a layer across
the electrode that becomes conductive only in those regions illuminated by the laser.
In a recent demonstration of the method,
the researchers used a 543-nm HeNe laser made by JDSU of Milpitas, Calif. They transmitted
the laser through the electrolyte and sample, resulting in a spot roughly 1 mm in
diameter. They positioned the beam — and, thus, the traps — with motor-controlled
mirrors driven by LabView software from National Instruments of Austin, Texas.
As detailed in the April 25 issue of
PNAS, James B. Harkins, Charles E. Witkowski and Dean G. Hafeman of Protein
Discovery used the setup to perform “dynamic steering” of a mixture
of proteins in an aqueous electrolyte. The researchers were assisted by Nathan
S. Lewis of California Institute of Technology.
First, they used the method to concentrate
the proteins, accumulating them from a 3-mm to a 1-mm spot. Then they moved the
beam of light in a counterclockwise direction, repositioning it in 2-mm steps every
five minutes. They observed the proteins moving with these steps as a single packet
of charged molecules; the proteins had been stained with different colors so that
the scientists could observe separation during this process.
They also showed that they could divide
a protein sample into two packets and could transport them independently with separate
high-field traps. After accumulating the sample into a smaller spot, they split
the laser beam into two illumination spots, each with the same intensity, duration
and diameter, spaced about 3 mm apart. They increased this distance by about 3 mm
every five minutes, and the packets of charged molecules followed.
The researchers noted that theoretical predictions
indicate that the charged molecules would lag behind the traps if they were
moved faster than 10 μm/s. Translating the molecules at a rate of 3 mm
per 5 min works because it is equal to this. In separate experiments, they confirmed
that the relative velocity of the molecules dropped precipitously when they moved
the light beam at a rate higher than 10 μm/s.
A photoconductive electrode
creates packets of charged molecules in the region illuminated by a laser, enabling
the transport of proteins and other smaller biomolecules that optical tweezers typically
cannot move without the aid of larger “handles.”
The technique could transport a variety
of biomolecules in addition to proteins; for example, oligonucleotides, DNA and
RNA. Therefore, it has potential for a number of applications, including the manufacture
of biochemical arrays, the transport, reaction and assembly of nanostructures and
other polymeric materials in microreaction volumes, and electrochromatographic separation.
It offers several advantages over conventional approaches to separation, which involve
gel electrophoresis and require high voltages and bulky equipment and which take
longer as the molecules move over very large distances.
But before the technique is ready for widespread application, the researchers must incorporate a relatively inexpensive photoconductive material as an electrode. Currently, they are using germanium, which
costs more than silicon. Materials such as titanium oxide would be considerably
less expensive, Thundat pointed out. They plan to test this material this summer.
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