posted on 2024-01-23, 23:46authored byTakahisa Matsuzaki, Ryuzo Kawamura, Akihisa Yamamoto, Hozumi Takahashi, Mai Fujii, Shodai Togo, Yosuke Yoneyama, Fumihiko Hakuno, Shin-Ichiro Takahashi, Masami Suganuma, Seiichiro Nakabayashi, Shivani Sharma, James K. Gimzewski, Hiroshi Y. Yoshikawa
Interference reflection microscopy (IRM) is a powerful,
label-free
technique to visualize the surface structure of biospecimens. However,
stray light outside a focal plane obscures the surface fine structures
beyond the diffraction limit (dxy ≈ 200 nm). Here, we developed an advanced interferometry
approach to visualize the surface fine structure of complex biospecimens,
ranging from protein assemblies to single cells. Compared to 2-D,
our unique 3-D structure illumination introduced to IRM enabled successful
visualization of fine structures and the dynamics of protein crystal
growth under lateral (dx‑y ≈ 110 nm) and axial (dx‑z ≤ 5 nm) resolutions and dynamical
adhesion of microtubule fiber networks with lateral resolution (dx‑y ≈ 120 nm),
10 times greater than unstructured IRM (dx‑y ≈ 1000 nm). Simultaneous reflection/fluorescence
imaging provides new physical fingerprints for studying complex biospecimens
and biological processes such as myogenic differentiation and highlights
the potential use of advanced interferometry to study key nanostructures
of complex biospecimens.