Imaging System Paves Way for Nanodevices

R&D DIGEST

Understanding the shape and structure of molecules in a biological nanomotor could lead to the development of nanodevices used for drug delivery and disease diagnosis. Researchers at Purdue University (West Lafayette, IN) have developed a sensitive imaging system that performs this job, providing a view of some small molecules, such as DNA and RNA.

The technology incorporates dual viewing of a single molecule using internal reflection and fluorescent imaging. It observes active molecules and detects their motion in real time. Key to the system is that it enables researchers to see the number of packaging RNA (pRNA) molecules contained in the DNA-packaging motor of the phi29 virus, a virus that infects bacteria. This is significant because identifying the shape of the motor and how it works could lead to the design of nanomotors that could contain and deliver DNA or RNA therapy to disease-causing cells.

Using a sensitive detection system and dual-color viewing, Purdue's technology differentiates and counts individual molecules within a nanodevice. Commercially available microscopes can't match this sensitivity, according to Peixuan Guo, PhD, professor of molecular virology and biomedical engineering at Purdue.

For example, Guo estimates that the phi29 motor is about 20 nm, and most microscopes only offer a resolution of 200 nm. And although some electron microscopes have the same resolution as the imaging system, the devices can't image RNA as well, because RNA does not have high electron density.

An assembly laser combiner provides the system with sensitivity. The combiner controls the release of the lasers, which serve as the light source. It also controls the delivery of the laser through optical fibers to the detector. To avoid confusing the detector, an advanced cooling system and a quartz prism are located atop the chamber, reducing background noise and light leakage.

The researchers determine the nanomotor's structure by looking at the interaction of the DNA and RNA. The molecules are tagged with red and green fluorescent labels that release signals. Before the signals reach the detector, they're divided based on color. Then a computer analyzes and converts them to images. The image indicates red and green dots for DNA and RNA, and yellow dots where the molecules overlap.

A graph containing lines is used to count the molecules. Through a process called photobleaching, a light shines on the fluorescent markers and ultimately destroys them. The graph drops as each molecule marker fades. Each stair step on the graph represents a molecule with one marker, explains Guo. The researchers counted the number of pRNA molecules and observed the biological motor's motion. From there, the device enabled the researchers to determine how many pRNA molecules were present in the biomotor. Guo says that such monitoring could be integral to diagnosis.

Purdue's research is funded by the National Institutes of Health. The team's work was published in a January issue of the European Molecular Biology Organization Journal.

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