How Do Your Molecules Size Up?

Protein Solutions' DynaPro molecular sizing instruments The scattering of sunlight by a combination of air molecules, dust particles, and water droplets creates some beautiful phenomena readily recognizable as white clouds, blue skies, and red sunsets. That scattered sunlight carries information that is detected by the eye and interpreted by the brain. In a similar fashion, laser light scattered by molecules in solution also carries valuable information. When properly detected and analyzed, for instance, this scattered light reveals much about a molecule's hydrodynamic radius, particle size and dimensions, polydispersity and molecular size distribution, molecular weight, concentration, and conformational changes. By using a technique called dynamic light scattering (DLS), these kinds of data are obtainable from several types of molecules and particles, including proteins, viruses, micelles, DNA, antibodies, lipids, and many more. The basic principles behind dynamic light scattering are straightforward. To begin with, a laser beam is directed into a sample of the molecule in an aqueous solution. When the particles are illuminated, the scattered light has a "speckle" pattern caused by interferences between the light scattered by individual particles. That is, a collection of bright and dark spots occurs as constructive and destructive interference spots form and interact. Because the particles move by Brownian motion, the intensity of the speckle pattern fluctuates as the particles move in relation to one another. This fluctuation in scattered light intensity has a time scale that is related to molecular size and is calculated by the mathematical technique of autocorrelation. Nonlinear curve-fitting of the function then provides information about the size distribution of the molecules. Because biomolecules scatter sufficient measurable light at low concentrations, the size and conformational properties of peptides, proteins, polysaccharides, and other supermolecular assemblies are easily studied. And since these studies are done in native solution conditions, aggregation phenomena and conformational changes as a function of temperature and physiological and nonphysiological conditions are also easily observed. Furthermore, DLS is a noninvasive technique so that samples may be recovered and then reanalyzed to monitor changes as a function of temperature, pH, ionic strength, sample concentration, and other parameters. Protein Solutions, Inc., of Charlottesville, Va., has applied DLS techniques to the development of its DynaPro family of molecular sizing instruments. Incorporating technology invented by Robert G.W. Brown, a pioneer in the light-scattering industry, with Protein Solutions' patented Avalanche Photodiode Detector Module, the DynaPro instruments are among the best dynamic light-scattering instruments available. Nancy Levinger, assistant professor of chemistry at Colorado State University, enthusiastically confirms this appraisal of the DynaPro instruments. "This is a great piece of equipment," she said during a telephone interview. "I researched four or five other companies that make this type of equipment before I purchased the DynaPro. The things that sold me on the DynaPro were its size--I can hold it in one hand--how fast it makes measurements, but most of all the fact that it sees very small particles. Most of the other manufacturers' equipment can't detect the small particles we measure. The extremely sensitive detection system that Protein Solutions uses, plus its sophisticated mathematics, allows the DynaPro to measure particles that other equipment can't." Protein Solutions' line of DynaPro instruments begins with the DynaPro Microsampler (MS). Its optical system features a removable 12µl quartz cuvette and precise engineering that guarantees perfect optical alignment for optimum sensitivity. While larger sample sizes are also available, the small sample sizes made possible by the 12µl cuvette are an attractive feature, especially for precious biological samples. If temperature studies are a consideration, Protein Solutions builds the DynaPro-MSTC that features a Peltier-based temperature-controlled cell. Capable of maintaining temperatures between 4°C and 40°C, the DynaPro-MSTC also reaches temperatures beyond this range with external waterbath connections. Of course, other features designed to enhance the sensitivity of the DynaPro are available, along with the DYNAMICS intuitive software package and the MicroFilter System for sample preparation. Applications for the DynaPro instruments abound, and generally fall into the broad categories of molecular characterization and conformation, interaction phenomena, and aggregation/self-assembly analysis. As examples, specific projects include identifying critical micelle concentrations of detergents and then determining the micelle size, studying the age effects on protein and peptide homogeneity, and predicting chances of successful crystallization by screening molecular size homogeneity prior to crystal growth experiments. The author, Michael Brush, can be contacted at mdbrush@compuserve.com. For additional information please contact Protein Solutions at (804) 817-7177, or visit the company Web site at www.protein-solutions.com.

How Do Your Molecules Size Up?

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