In talks delivered during the March 28-April 1 at the American Chemical Society annual meeting in Anaheim, researchers from Duke's Pratt School of Engineering also told how they are using an atomic force microscope to create reprogrammable "nanopatterns" of large biologically-based molecules that could potentially serve to analyze the protein contents of individual cells, among other uses.
The molecules are reprogrammable in the sense that they could be activated, deactivated, and then activated again for another use. They could serve as analytical tools because they could capture and isolate proteins of interest from complex mixtures.
The molecular dimensions of this work -- at the billionths-of-a-meter scale ("nano" means billionths) -- "introduces the concept of scaling-down chemistries to very small lengths," said Stefan Zauscher, a Duke assistant professor of mechanical engineering and materials science.
Zauscher was an organizer of a society symposium called "Smart Polymers on Colloids and Surfaces. "Smart" polymers are long-chained molecules that can reversibly change their conformations as well as reversibly and selectively bind to other molecules, Zaucher explained in an interview.
Besides nanobrushes, other examples of "smart" large molecules include those that interact through molecular recognition, such as streptavidin and biotin, and the biologically inspired elastin-like polypeptides (ELPs). Duke engineering researchers have developed ways to pattern all these constituents so they can react at nanoscale dimensions, he said. "One reason is simply the challenge: can we make features this small? Also, making features that small means you could get away with using very small amounts of chemicals, for example of proteins you might want to detect."
Zaucher, associate professor of biomedical engineering Ashutosh Chilkoti and other Pratt School investigators can now lay down patterns of the molecule streptavidin with dimensions of a few hundred nanometers. They do this with a special "dip pen" technique developed at Northwestern University that lets them turn an atomic force microscope (AFM) into a nanoscale quill. An AFM is a microscope that can image surfaces and detect forces at resolutions approaching the atomic scale.
Steptavidin, a natural protein that comes from the Streptomyces bacterium, can bind especially tightly with biotin, which is a member of the vitamin H family that occurs widely in nature. The nanopatterned streptavidin can thus link with other proteins that have been treated with biotin. This arrangement is thus potentially useful for grabbing proteins of interest out of large multi-molecular mixtures for analysis, Zauscher said.
By substituting similarly binding iminobiotin for biotin, the investigators can also remove the chemicals from a surface and start again, he said – in essence like erasing a blackboard. That's because iminobiotin, unlike biotin, releases its binding grip when the solution becomes highly acidic.
The Duke investigators used the same AFM method to deposit patterns as small as 200 nanometers of elastin-like polypeptides (ELPs). These protein-like molecules, which are similar to elastins in animal connective tissue, were genetically engineered in Chilkoti's laboratory.
The immobilized ELPs can then reversibly bind with other ELP-tagged proteins in solution, allowing them to potentially select out individual molecules for identification. Such a tiny ELP array might be used, for example, to screen the protein contents of an individual cell, Zauscher said.
A paper in the February 2004 issue of the research journal Nano Letters describes how Zauscher's group has fabricated patterned nanobrushes using what is called the atom transfer radical polymerization method. This method allows certain polymers to grow from a surface in controlled reactions that makes them sprout in brush-like shapes between 30-500 nanometers long.
An AFM was initially used to lay out nanopatterns from which the bristles grow in what he called a "pseudo-living" manner, but not by the dip pen technique. Instead, the AFM's tip was used like a back hoe, Zauscher said, to "nanoshave" channels for the initiating chemical. The Duke researchers found beams of electrons can also be used to create the nanopatterns in lieu of an AFM.
The "smart" polymer used is stimulus-responsive, in that its long chained brushes can change their conformation by adding solvents or changing the temperature.
Such nanobrushes could potentially be used as repeat-use chemical detectors in tiny "microfluidic" devices such as a lab on a microchip, he said.
Journal
Nano Letters