Optical Detection Of Photorelease Kinetics On Gold And Glass Surfaces Using Streptavidin-Coupled Biotinylated Photolabile Protecting Groups For Nucleosides

Five biotinylated photolabile compounds of the general structure Bt-L-1-NPPOC-X-L-2 were synthesized, in which Bt represents a biotin unit, L-1 is a 3,6-dioxa-n-octane or an n-hexane spacer, NPPOC is the photolabile protecting group 2-(2-nitrophenyl)propoxycarbonyl, and X is a thymidine unit as a representative nucleoside or a direct linkage to L-2, an -mercapto- or -aminohexoyl linker, for coupling to a substrate surface. These compounds served for testing the photocleavage kinetics in self-assembled monolayers on gold or glass by using surface plasmon resonance (SPR) on gold or reflectometric interference spectroscopy (RIfS) on glass, whereby the biotin moiety offered the possibility to increase the bulkiness of the leaving group by binding to streptavidin, which thereby largely enhanced the SPR or RIfS signals. The photokinetics, found to consist in a dominating fast stage and a less contributing slow stage, were quantitatively analyzed, and the quantum yield of the fast part reached values up to almost 1 in favorable cases. A direct comparison of the results from SPR and RIfS yielded almost identical results. The present investigations pave the way to insitu monitoring of the photolithographic synthesis of DNA chips.