Electron Transfer Dissociation
Mass spectrometry has become the method of choice for protein analysis and biomarker discovery. At “Pittcon 2007”, Thermo Scientific introduced electron transfer dissociation (ETD) on a linear ion trap mass spectrometer. ETD is a new method that fragments proteins in order to pinpoint important system biology changes in structure that occur in response to a growth factor or other stimulation.
ETD is available on the Thermo Scientific LTQ XL™ mass spectrometer. “ETD is a revolution for proteomic analysis because it dramatically increases the amount of sequence information you can get from larger proteins,” says Ian Jardine, Ph.D., vp of global R&D at Thermo Fisher Scientific. ETD works best with linear ion traps that Thermo Scientific provides for the LTQ XL.
ETD, which improves on the traditional method of collision-induced dissociation (CID), provides important new peptide structure information not available from conventional dissociation methods, such as post-translational phosphorylation, according to Dr. Jardine. He predicts that in two years, every high-end proteomics lab will be using ETD to stay competitive, and in three to five years, ETD will be used routinely in all proteomics labs. “It is too powerful a tool not to use,” adds Dr. Jardine.
Systems biology researchers use mass spectrometry for SILAC (stable isotope labeling of amino acids in cell culture) experiments. Using the older CID method, researchers at the Max Planck Institute detected 2,000 proteins and 6,000 phosphorylation sites after adding epidermal growth factor to cells. If ETD were added to this type of systems biology experiment, the number of proteins and phosphorylation sites would increase dramatically, Dr. Jardine says. “ETD moves information about cells to the next level.”
Because few companies offer cell culture media and isotope reagents to perform SILAC experiments, Thermo Scientific rolled out a full suite of SILAC kits at the “American Society for Mass Spectrometry meeting” in June. “A good mass spectrometer to do readouts of SILAC is the LTQ,” says Dr. Jardine, “and now we have the reagents to seamlessly carry out experiments.”
Thermo Scientific’s Dharmacon siRNA product line includes kits that cover the entire human and mouse genome. Although these first-generation siRNA products show good specificity, they also activate unintended genes, Dr. Jardine says. “Dharmacon’s new On-Target Plus™ products greatly reduce off-target problems.”
At the University of Texas Southwestern Medical Center in Dallas, researchers recently made a groundbreaking discovery using Thermo Scientific Dharmacon siARRAY® Whole Human Genome collection of siRNA molecules. From among 21,000 genes, they identified 87 genes that increase the susceptibility of human cancer cells to the drug paclitaxel (Taxol). When these genes were blocked, paclitaxel is up to 10,000 times more effective, as described in an April 2007 issue of Nature. The study suggests that by silencing key genes, patients can be treated with lower doses of toxic chemotherapy drugs.
The future looks bright for Thermo Fisher’s siRNA products, Dr. Jardine explains, but problems remain to be solved. One is that siRNA reagents cannot enter all cells.
“We need to solve that problem to expand its utility,” says Dr. Jardine. Moreover, it needs to be seen whether siRNA silences genes in the body. “That’s the next level of research,” he adds.