Photophysical properties and synthesis of new dye-cyclooctyne conjugates for multicolor and advanced microscopy.
Cyclooctyne conjugates with fluorophores are often used for bioorthogonal labeling in cells and tissues. However, no comprehensive library of one cyclooctyne core structure with different fluorescent dyes spanning the whole visible spectrum up to the NIR had been described so far. Hence, we synthesized and evaluated one cyclooctyne core structure which is easily accessible for the attachment of different dyes for multicolor imaging, FRET analysis, and study of metabolism in vivo. For these reasons we developed an easy one step synthesis starting from a known cyclooctyne. In combination with NHS-activated dyes, the cyclooctyne reacted to the dye DAB-MFCO conjugates within only 1-2 h at room temperature with high yields. We created conjugates with dyes that have high brightness and are bleaching stable with wavelengths from green to NIR. The ability to label glycans on cell surfaces was tested. All dye DAB-MFCO conjugates undergo click reactions on azide functionalized glycan structures with satisfactory photophysical properties. In total, seven different dye DAB-MFCO conjugates were synthesized; their photophysical properties and suitability for click labeling in biological applications were evaluated, making them suitable for single molecule and high resolution measurements.
†Institute of Organic Chemistry, Karlsruhe Institute of Technology, Fritz-Haber-Weg 6, 76131 Karlsruhe, Germany.|‡Light Technology Institute, Karlsruhe Institute of Technology, Engesserstraße 13, 76131 Karlsruhe, Germany.
Anna
A
Hörner
†Institute of Organic Chemistry, Karlsruhe Institute of Technology, Fritz-Haber-Weg 6, 76131 Karlsruhe, Germany.
Tobias
T
Hagendorn
†Institute of Organic Chemistry, Karlsruhe Institute of Technology, Fritz-Haber-Weg 6, 76131 Karlsruhe, Germany.|§Institute of Toxicology and Genetics, Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen, Germany.
Ute
U
Schepers
†Institute of Organic Chemistry, Karlsruhe Institute of Technology, Fritz-Haber-Weg 6, 76131 Karlsruhe, Germany.|§Institute of Toxicology and Genetics, Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen, Germany.
Stefan
S
Bräse
D000480:Alkynes / Q000737:chemistry*; D001386:Azides / Q000737:chemistry*; D057930:Click Chemistry; D003300:Copper / Q000737:chemistry; D003500:Cyclization; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000737:chemistry*; D054879:Halogenation; D006367:HeLa Cells; D006801:Humans; D007259:Infrared Rays; D008853:Microscopy / Q000379:methods*; D011134:Polysaccharides / Q000032:analysis / Q000737:chemistry*; D011700:Putrescine / Q000737:chemistry
D000480:Alkynes; D001386:Azides; D005456:Fluorescent Dyes; D011134:Polysaccharides; D003300:Copper; D011700:Putrescine