The Odyssey Fc imager is great in terms of ease of use and quality of data, and also helps us conserve bench space.
Dr. Rick Page
Miami University in Oxford, OH
At Miami University in Oxford, OH, assistant professor Dr. Rick Page is unfolding the fate of misfolded membrane proteins. Page and his research team study protein quality control, and how defects in protein conformation result in disease.
In response to a misfolded protein, cells either recruit the refolding machinery via heat-shock protein 70 (Hsp70) or initiate a protease cascade through the C-terminus of Hsp70 Interacting Protein (CHIP) to degrade that protein. "In cystic fibrosis (CF) for instance, a mutation causes the cystic fibrosis transmembrane regulator (CFTR) to fold at a slow pace. Cells recognize this as a folding defect and recruit CHIP to degrade CFTR prematurely. As a result, there is not sufficient CFTR at the surface of cells in CF patients," says Page. Their group has identified that the Receptor for Activated C kinase 1 (RACK1) protein affects the surface regulation of CFTR, through its interaction with filamin-A1.
In order to isolate proteins, elucidate their structures, and characterize their functions, Page’s team employs various biochemical assays coupled with biophysical techniques like X-ray crystallography and NMR studies. The Odyssey® Fc Imaging System housed in the lab has found utility as the go-to device for all of their gel imaging needs1,2,3.
Convenient, Versatile Gel Imaging
"We image our ethidium bromide stained agarose gels for DNA work on the Odyssey Fc system. We also stain SDS-PAGE gels with Coomassie, and use the 700 nm channel for detecting the protein of interest in eluted chromatography fractions," says Page. Additionally, the group performs fluorescent Western blotting, especially ubiquitin assays on the Odyssey Fc imager.
"CHIP can ubiquitinate Hsp70. One of the reasons we really like using the Odyssey Fc system is that we can probe ubiquitinated vs. non-ubiquitinated Hsp70 at the same time. The [Odyssey Fc] imager has the resolution that we need to tell the bands apart," he adds.
Instead of getting two or three different instruments for imaging Western blots, agarose gels, SDS-PAGE gels, and other assays, we purchased a single instrument that can be used for all of these applications.
Page emphasizes that they don’t have a large lab space, and hence bench space is at a premium. "The footprint of the Odyssey Fc system is not too large, which is good. Instead of getting two or three different instruments for imaging Western blots, agarose gels, SDS-PAGE gels, and other assays, we purchased a single instrument that can be used for all of these applications. The Odyssey Fc imager is great in terms of ease of use and quality of data, and also helps us conserve bench space."
Simplicity and Reliability with Infrared Detection
The Odyssey Fc in Page’s lab was purchased through the LI-COR® Science Undergraduate Research Grant (SURG) program. Page teaches a course in biophysics wherein students learn about the workings of the Odyssey Fc system and gain hands-on experience using it in the research lab. "Students actually use the Odyssey Fc imager themselves; a few have performed quantification as well. I have even had freshmen students image their gels and Western blots on the imager. The instrument is simple enough to use, and once trained they can work independently," he says.
Page asserts that film-based chemiluminescent Western blot detection depends on exposure and other confounding factors. "We wanted a better way of imaging Western blots than using film. We tried fluorescence detection, and it worked right off the bat. The Odyssey Fc imager has an incredibly high resolution and gives you a lot of customization. I learned to love the system."
Academic scientists like Page are bringing reliability and convenience to their research and classrooms with the user-friendly Odyssey Fc technology.
Publications resulting from work on the Odyssey CLx
- Smith, L., Litman, P., Kohli, E., Amick, J., Page, R. C., Misra, S., & Liedtke, C. M. (2013). RACK1 interacts with filamin-A to regulate plasma membrane levels of the cystic fibrosis transmembrane conductance regulator. American Journal of Physiology. Cell Physiology, 305(1), C111–20. doi:10.1152/ajpcell.00026.2013
- Page, R. C., Pruneda, J. N., Amick, J., Klevit, R. E., & Misra, S. (2012). Structural insights into the conformation and oligomerization of E2~ubiquitin conjugates. Biochemistry, 51(20), 4175–4187. doi:10.1021/bi300058m
- Amick, J., Schlanger, S. E., Wachnowsky, C., Moseng, M. A., Emerson, C. C., Dare, M., et al. (2014). Crystal structure of the nucleotide-binding domain of mortalin, the mitochondrial Hsp70 chaperone. Protein Science, 23(6), 833–842. doi:10.1002/pro.2466
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