Design of fluorescent dyes for biomedical applications

Search and development of novel highly efficient fluorescent probes for biology and medicine are among the main activities of our group. The "lead dye" method, an original approach to design of fluorescent probes with required properties, was proposed and applied by us for the performing of contract research projects. The wide house-stock collection containing about 2,000 dyes of various classes (polymethine cyanines, styryles, coumarines, and metallocomplexes) is used as a basis for performing of such studies.

In collaboration with BioRad Inc. we developed squaraine dyes for using as unspecific fluorescent probes for detection of proteins in separation systems.

Series of sensitive fluorescent dyes for nonspecific detection of proteins in gels (LUCY dyes) was developed by our group for Sigma-Aldrich Inc. Main advantages of the stains Lucy 506, Lucy 569 and Lucy 565 are their high sensitivity, simple and rapid staining protocol and low protein-to-protein variability. These dyes show a wide linear concentration range and allow detection of about 5-10 ng of protein per band (Fig. 6).

Figure 6. Limits of proteins detection on SDS-polyacrylamide gel by LUCY-506 and LUCY-565 proteins gel stains developed by us for Sigma-Aldrich Inc., in comparison with SYPRO Ruby stain (Invitrogen Corp.).

In collaboration with Sigma-Aldrich we successfully developed novel high-sensitive fluorescent probe Nancy-520 for DNA visualization in gels. With this stain one can visualize as low as 0.5 ng/band of dsDNA in agarose gel. Besides, Nancy-520 can be used to determine dsDNA concentrations in solution from 0.15 µg/ml DNA.

• W. Patton, S. M. Yarmoluk, P.Pande, V.B. Kovalska, D.Lijun, K.Volkova, J. Coleman, M.Losytskyy, A.Ludlum, A.Balanda/ Patent Application Publication. Dyes for analysis of protein aggregation WO 2011/065980 A2, 03.06.2011.
• A. Rueck, B. Schoenenberger, S. Yarmoluk, V. Kovalska, M. Losytskyy, Y. Slominskii. Nucleic acid fluorescent stains // US Patent Application Publication US2010041045, 18.02.2010.
• T.R. Berkelman, S.M. Yarmoluk, V.B. Kovalska, M.Yu. Losytskyy, K.D. Volkova. Use of squaraine dyes to visualize protein during separations // Patent Application Publication WO 2008/027821 A1, 06.03.2008.
• K.D. Volkova, V.B. Kovalska and S.M. Yarmoluk. Modern techniques for protein detection on polyacrylamide gels: problems arising from the use of dyes of undisclosed structures for scientific purposes // Biotech. Histochem. – 2007. – Vol. 82, Iss. 4&5. – P.201-208.
• V. Kovalska, D. Kryvorotenko, M. Losytskyy, P. Nording, A. Rueck, B. Schoenenberger, S. Yarmoluk, F. Wahl. Detection of polyamino acids using trimethincyanine dyes // US Patent Application Publication US2006207881, 21.09.2006.