Chen et al. used GLAD to fabricate a patterned substrate. To improve the hotspots and enhancement, plasmonic gratings with periodic nanogaps were developed. Silver film growth on polymethylsilsesquioxane (PMSQ) polymer gratings were replicated from HD DVD gratings molds. The silver was deposited at an angle of α = 60 using GLAD. SM fluorescence images were taken on these GLAD-based gratings for a DNA/RNA duplex tagged with cyanine 3 and cyanine 5 labels. GLAD gratings with incorporated nanoprotrusions enabled SM imaging over a wide range of fluorescence from 50 pM to 10 uM. Additionally, the nanoprotrusions allowed the group to use multiple-emitter fitting analysis to solve the crowded field problem created by higher fluorophore concentration. Patchy microspheres fabricated using GLAD setup have also shown potential application in improving the diffraction limit of super-resolution microscopy [221]. Wood et al. performed similar work whereby they used DVD to produce gratings in polymethylsilsesquioxane (PMSSQ) deposited on a glass substrate via microcontact lithography. Ag was deposited using GLAD fabrication. This formed nanoprotrusions and nanogaps that enhanced the LSPR activity and also increased the hotspots, exciting the fluorophores [222].
Shang et al. fabricated microsphere-nanospheres by coating 100 nm thick Ag films using GLAD on the BaTiO3 glass (α = 60), and the patchy microsphere was made from p-BTG particles. The imaging contrast of p-BTG was found to be improved by a factor of 6.5. Overall, this patchy microsphere improved the imaging contrast without the use of immersion liquid and led to greater advances in the field of super-resolution microscopy.
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