Improving the Safety and Sensitivity While Speeding Up the Process
Electrophoresis relies on a basic process— particles moving in an electric field, more or less. Although known for more than 200 years, this phenomenon still drives fundamental techniques in many laboratories. Despite that long history, this field still revolves around time. Instead of centuries, though, researchers now focus on minutes. In particular, they want the process to take less time—much less. On the other hand, everyone also wants more accuracy and lower detection limits, all with higher throughput.
When asked about the biggest challenges today in gel electrophoresis, Nikita Warner, senior product manager at Life Technologies (Carlsbad, CA), says quickly, “Time to results. The desire is always faster and faster.” She points out that running nucleic acids on a gel accelerated from 45 minutes to 30 minutes about a decade ago, and then dropped to just 10 minutes. “With real-time visualization, you can stop a run as soon as you see the band, and that can be even less than 10 minutes,” she says.
The long history of gel electrophoresis plays some role in the widespread use of this technology. Kevin Dorfman, Ph.D., associate professor of chemical engineering and materials science at the University of Minnesota, points out that gel electrophoresis for DNA separations is “pretty well-established technology.” As he says, “As long as your separation is good enough, you can just cut out the bands.” It’s really as simple as taking a scalpel and literally cutting out the bands of interest. Still, advances in technology can improve this form of electrophoresis. As an example, Dorfman notes that some commercially available buffers for the gels don’t heat up as much as traditional ones do, and that speeds up a separation.
The old and the new
Although people are programmed to see things digitally these days, says Adam Sartiel, senior manager, business and product development at Life Technologies Israel, “electrophoresis is still pretty old-school, because it provides analog results that must be transformed to a digital format.” Even when a platform provides a digital output, users still want to see the bands—the analog version of the data. An experienced researcher might find that the analog information helps, especially with any troubleshooting.
Being old-school with electrophoresis actually starts before running any gels. It begins with making them. “Most people still use manual methods to prepare gels,” says Sartiel, “and that’s not precise, not a calibrated system.”
Beyond consistency, safety also plays a role in making gels. With regard to polyacrylamide gel electrophoresis (PAGE), Warner points out the danger of handling toxic chemicals. “With agarose gels,” she adds, “the personal hazard is not as great.”
Consequently, the Life Technologies experts see more people turning to commercially prepared gels. In addition to being more consistent and safer for users, Warner says their gels provide much longer shelf life. “When you make your own gels for PAGE, they only last several hours at best,” she says. As an example, she points out that the NuPAGE gels have a yearlong shelf life or more.
New uses for an old technology
Some new technologies increase the use of gel electrophoresis, and next-generation sequencing provides a crucial example. “Some sequencing systems require the ability to extract libraries of a certain size from a gel, and that can now be done quickly and automatically with gel electrophoresis,” says Sartiel.
Variations in electrophoresis, such as capillary electrophoresis, also impact the potential applications. As Mark Lies, Ph.D., global manager for capillary electrophoresis at Beckman Coulter (Brea, CA), says, “The separation format lets you control the environment inside the capillary, allowing you to run back-to-back separations on completely different types of molecules using different assays.”
According to Lies, capillary electrophoresis can be applied to solve many problems. “It can be used to assess purity and charge heterogeneity of proteins or to analyze the carbohydrates associated with those proteins,” he says. Lies also points out that capillary electrophoresis can be used to analyze ion populations in a sample or to separate enantiomers from one another. He adds, “There’s a very strong adoption of capillary electrophoresis in the biopharmaceutical industry for automated, quantitative characterization of therapeutic proteins. Biotechnology companies are counting on [capillary electrophoresis] technology to help maintain quality control of their development and commercialized monoclonal-antibody therapies.” He also sees this technology used for a range of applications in the petrochemical, environmental, food and beverage, and biofuel industries.
If a researcher wants to identify the components of a mixture of DNA with quantitative precision, Dorfman recommends capillary electrophoresis. On the other hand, he mentions that a researcher can set up gel electrophoresis with a much smaller investment.
“Capillary electrophoresis provides automated analysis so user variability is minimized,” Lies says. “In addition, integrated software and electronic technical controls provide precise instrument control, data quantification, and the ability to comply with regulatory requirements.”
Besides the obvious advantage of decreasing variability, automated operation provides method-development capabilities in which a user can program the instrument to alter separation conditions. “Integrated software lets you control the instrument,” Lies explains. For example, he says, “The voltage applied to create the field strength and the temperature of the separation may impact the separation.”
It’s even possible to add a sample preparation platform to the front end of a capillary-electrophoresis platform. “Then you gain run-to-run reproducibility because there is minimal manual interaction in the sample prep process too.”
Overall, the growing capabilities of capillary electrophoresis give users more separation options with high precision, accuracy, and robustness. Consequently, this technology will begin to apply to more types of analysis in a growing number of fields. In addition, improvements in the components of this centuries-old concept and completely new technologies drive this field ahead.