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The answers to big questions – how to create green fuels or how to cure deadly diseases – may have their answers in tiny motions in living cells. To understand these small movements, Ning Fang, associate scientist, is using optical microscopy to see how nanoparticles are manipulated by “nanomachines” in cells.

In living cells, numerous biological nanomachines perform various functions.  But according to Fang, scientists have only a limited understanding of how these nanomachines work, especially in cellular environments. And because the malfunction of any of these nanomachines can lead to diseases, there is a great need for new techniques to help investigate the composition, dynamics and working mechanisms of these nanomachines. Nanomachines are also very efficient at converting chemical or other energy into motion, so understanding them may give scientists ideas for how to build efficient green-energy technology at the micro- or macroscale.

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Ning Fang is using optical microscopy to study the mechanisms of biological nanomachines. Here,
a gold nanorod is attached to a microtubule. Tracking the movement of the nanorod reveals the
rotation of the microtubule

To understand how these nanomachines work, scientists examine how nanoparticles move with various types of motion induced by nanomachines that are essential to their function.

Translational motion, or movement in which the position of an object is changed, can be tracked through a variety of current techniques.  However, rotational motion, which is as important and fundamental as translational motion, was largely unknown due to technical limitations.

Previous techniques, such as particle-tracking or single-molecule fluorescence polariz-

 

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Left: Transmission electron microscopy image of gold nanoparticles. Right: Differential
interference contrast images of two 25 by 73 nanometer gold nanorods positioned at different
orientations. The periodic changes of bright and dark intensities allows the nanorods’ orientation
to be determined with high precision.

ation, only allowed rotational movement to be resolved in simplified, controlled in vitro systems.  In its research, Fang’s group has gone beyond studying motions in the in vitro environment to imaging rotational movement in the in vivo, or live cell, environment.

To do this, the group relies upon the use of gold nanorods, which are only 25 by 73 nanometers in size (a well-packed bundle of 1000 nanorods has the same diameter as a human hair).  In live cells, these nontoxic nanorods scatter light differently depending upon their orientation.  Using a technique called differential interference contrast microscopy, or DIC, Fang’s team can capture both the orientation and the position of the gold nanorods in addition to the optical image of the cell and, thus, reveal a particle’s 6D (three spatial coordinates, two orientation angles and time) movement within living cells.

“This new technique opens doors to understanding the working mechanism of living nanomachines by revealing their complex internal motions,” says Fang. “Studying rotational motions at the nanometer scale inside a living cell is just something that has never been done before.”  

He added that understanding this rotational motion is important in the fight against diseases, such as Alzheimer’s, because it can help scientists better understand how neurons are impacted by the environment. 

Fang’s research group is also

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(from left) Ning Fang, Wei Sun and Gufeng Wang

developing another type of DIC microscopy that generates a high-contrast, high-resolution view of materials that would be invisible under conventional light microscopes, so Fang and his colleagues can visualize the composition of block copolymers.

“DIC uses a special principle based on interference so we can see inside these polymers down to about a domain of a few tens of nanometers or even smaller,” Fang says. “The polymers appear to have different patterns of nanostructures, and we are now hoping that this understanding will lead to applications in clean energy.”

In addition, Fang’s team is working in the area of super-resolution fluorescence imaging, with the goal of visualizing ever finer details of materials.

“The overall goal is to break the diffraction limit of light,” says Fang. “For example, we want to be able to distinguish between two fluorescent molecules that are so close to each other that they previously would have appeared as one.”

His team’s technique, called 3D super-resolution fluorescence microscopy, has the added benefit of automation.

“Compared to others’ systems, we can control most parts of the microscope by computer, so the computer automatically calibrates and carries out the scan,” says Fang. “This means the system is very convenient for end users, and, more importantly, we can better reproduce the results of our imaging process.”

In all these types of imaging, Fang’s group is working to improve the ability to manipulate samples during imaging.

“We need better control of the sample so that we can pinpoint when and how we introduce external triggers to samples,” explains Fang. “For instance, we are studying the enzymatic reaction for breaking down cell walls to examine how to best make biofuel. That type of experiment requires introducing a reagent to the sample to start the reaction.”

The controls are provided by a cleverly designed microchannel network, and Fang and his colleagues have introduced the first type that can work with high-fidelity optical imaging.

Overall, Fang’s goal is to improve on the complete platform of optical imaging.

“We seek to improve localization, that is, we want to see a particle’s location with nanometer precision, and we want to improve resolution, so we can identify two close particles as individuals,” says Fang. “We also want to be able to track particles’ movements, and we want to be able to manipulate the samples.”

“The complete platform of imaging, combined with collaborating with others at Ames Laboratory with different techniques, is one advantage we have here at Ames Lab,” adds Fang. “We can use all of our techniques to examine one material. We gain more basic understanding of the material that may someday lead to important applications.”

~ by Breehan Gerleman Lucchesi