RESUMO
Fluorescence-based expansion microscopy (ExM) is a new technique which can yield nanoscale resolution of biological specimen on a conventional fluorescence microscope through physical sample expansion up to 20 times its original dimensions while preserving structural information. It however inherits known issues of fluorescence microscopy such as photostability and multiplexing capabilities, as well as an ExM-specific issue in signal intensity reduction due to a dilution effect after expansion. To address these issues, we propose using antigen-targeting plasmonic nanoparticle labels which can be imaged using surface-enhanced Raman scattering spectroscopy (SERS) and dark-field spectroscopy. We demonstrate that the nanoparticles enable multimodal imaging: bright-field, dark-field and SERS, with excellent photostability, contrast enhancement and brightness.
RESUMO
Single-molecule Surface-Enhanced Raman Scattering (SERS) detection of buckminsterfullerene (C(60)) is achieved by using different isotopologues of the molecule with a distribution around an average isotopic substitution ((12)C â (13)C) of ~30%. The distribution of different isotopologues creates a broad (~20 cm(-1)) average SERS signal within which single-molecule SERS spectra of individual isotopic realizations of the molecule can be distinguished. The SERS enhancement factors for SM-SERS C(60) events are typically in the range of ~10(8), suggesting a limitation imposed by either photobleaching or surface interactions with the (Ag) metallic colloids to reach the highest SERS hot-spots (which can typically have larger maximum enhancements). SM-SERS signals of isotopically substituted C(60) also show broader peaks (FWHM ≈ 4 cm(-1)) than equivalent signals in natural C(60). The latter feature suggests a contribution to the homogeneous broadening coming from isotopic disorder in the molecule; a feature that can only be observed with the ability to detect single-molecule spectra.
