The new technique, developed by a research team led by University of Illinois at Chicago assistant professor of chemistry Lawrence Miller, provides the clearest, most dynamic view yet of protein-protein interactions in cells when viewed through a specially modified microscope.
The finding is reported in the Proceedings of the National Academy of Sciences (advanced online July 19.)
Knowing where and when particular proteins interact within the cell is key to understanding life processes at the molecular level.
In a technique called luminescence resonance energy transfer, two proteins in a cell are labeled with differently colored, luminescent molecules that absorb light of one color and give it off as another color. By taking several pictures of the cell and mathematically analyzing the pictures, researchers gain information about the proteins' precise location and whether they are interacting.
Miller and his team used a novel type of luminescent molecule for labeling, making it possible to get the same information using fewer pictures. This simplifies the analysis and allows for five-fold faster data acquisition. Images show a 50-fold improvement in sensitivity.
Working with Jerrold Turner, professor and associate head of pathology at the University of Chicago, Miller used a hybrid chemical/genetic approach to tag the proteins of interest. One of the proteins was genetically modified so that it would bind to a terbium complex. The terbium complex has an unusually long time between light absorption and emission. The second target protein was genetically modified to link to a fluorescent tag with a short emission lifetime. When the two proteins interact, the luminescent tags are brought very close together, generating a unique luminescent signal that can be seen under a microscope.
Miller and his colleagues modified a conventional microscope to exploit the long lifetime of the terbium protein tags. Pulsed light is used to trigger the terbium luminescence, detected after the other luminescent species within cells have gone dark, allowing unwanted background to be removed from the image.
The new technique "increases sensitivity and makes the whole process faster," Miller said. "This increases the time-resolution of the experiment, allowing you to see how interactions change on a faster time scale, which can help to better figure out how certain biological phenomena work."
The technique required a reliable way to deliver the luminescent terbium probe through a living cell membrane without contaminating or damaging the cell. The researchers developed a way to co-opt pinocytosis, the process by which cells drink in small amounts of surrounding fluid.
"With this new tool, we hope cell biologists and others will be able to study things they haven't seen before, such as interactions that couldn't be visualized in live cells in real time," Miller said. "Hopefully the method will yield information that makes it easier to deduce biological mechanisms."
Other authors include UIC graduate students Harsha Rajapakse (the lead author), Nivriti Gahlaut and Shabnam Mohandessi and University of Chicago graduate student Dan Yu. The terbium tag was developed in collaboration with Richmond, Calif.-based Lumiphore, Inc. Major funding was provided by the National Institutes of Health and the Chicago Biomedical Consortium.
Paul Francuch | Newswise Science News
Transport of molecular motors into cilia
28.03.2017 | Aarhus University
Asian dust providing key nutrients for California's giant sequoias
28.03.2017 | University of California - Riverside
The Institute of Semiconductor Technology and the Institute of Physical and Theoretical Chemistry, both members of the Laboratory for Emerging Nanometrology (LENA), at Technische Universität Braunschweig are partners in a new European research project entitled ChipScope, which aims to develop a completely new and extremely small optical microscope capable of observing the interior of living cells in real time. A consortium of 7 partners from 5 countries will tackle this issue with very ambitious objectives during a four-year research program.
To demonstrate the usefulness of this new scientific tool, at the end of the project the developed chip-sized microscope will be used to observe in real-time...
Astronomers from Bonn and Tautenburg in Thuringia (Germany) used the 100-m radio telescope at Effelsberg to observe several galaxy clusters. At the edges of these large accumulations of dark matter, stellar systems (galaxies), hot gas, and charged particles, they found magnetic fields that are exceptionally ordered over distances of many million light years. This makes them the most extended magnetic fields in the universe known so far.
The results will be published on March 22 in the journal „Astronomy & Astrophysics“.
Galaxy clusters are the largest gravitationally bound structures in the universe. With a typical extent of about 10 million light years, i.e. 100 times the...
Researchers at the Goethe University Frankfurt, together with partners from the University of Tübingen in Germany and Queen Mary University as well as Francis Crick Institute from London (UK) have developed a novel technology to decipher the secret ubiquitin code.
Ubiquitin is a small protein that can be linked to other cellular proteins, thereby controlling and modulating their functions. The attachment occurs in many...
In the eternal search for next generation high-efficiency solar cells and LEDs, scientists at Los Alamos National Laboratory and their partners are creating...
Silicon nanosheets are thin, two-dimensional layers with exceptional optoelectronic properties very similar to those of graphene. Albeit, the nanosheets are less stable. Now researchers at the Technical University of Munich (TUM) have, for the first time ever, produced a composite material combining silicon nanosheets and a polymer that is both UV-resistant and easy to process. This brings the scientists a significant step closer to industrial applications like flexible displays and photosensors.
Silicon nanosheets are thin, two-dimensional layers with exceptional optoelectronic properties very similar to those of graphene. Albeit, the nanosheets are...
20.03.2017 | Event News
14.03.2017 | Event News
07.03.2017 | Event News
28.03.2017 | Life Sciences
28.03.2017 | Information Technology
28.03.2017 | Physics and Astronomy