FINDING FLUORESCENCE POLARIZATION ANISOTROPY FROM DIGITAL MICROIMAGES OF CELLS
I. G. Pal’chikova1,2, L. V. Omel’yanchuk3, N. V. Kamanina4, S. N. Makarov1,2, E. S. Smirnov1,2
1Technological Design Institute of Scientific Instrument Engineering, ul. Russkaya 41, Novosibirsk 630058 Russia
palchikova@tdisie.nsc.ru
2Novosibirsk State University, ul. Pirogova 2, Novosibirsk, 630090 Russia
3Institute of Molecular and Cell Biology, Siberian Branch of the Russian Academy of Sciences, pr. Akademika Lavrent’eva 10, Novosibirsk, 630090 Russia
ome@mcb.nsc.ru
4Vavilov State Optical Institute, Birzhevaya Liniya 12, St. Petersburg, 199034 Russia
nvkamanina@mail.ru
Keywords: digital microimage processing, fluorescence polarization anisotropy, homo-FRET, proteins, GFP-labeled
Abstract
Methods are developed to record and process microimages which are suitable to obtain quantitative information about the fluorescence polarization anisotropy caused by resonance energy transport between green fluorescent protein (GFP)-labeled proteins in a living cell. The methods allow for accurate detection of dimers and higher order associates for GFP-labeled proteins. Protein-protein interaction between subunits of the trimeric GFP protein was recorded. The sources of hardware inaccuracies were found and ways to eliminate them were proposed, which reduced the coefficients of data variation by an order of magnitude in comparison with the previously obtained values.
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