Quantitative multicolor subdiffraction imaging of bacterial protein ultrastructures in three dimensions.

Gahlmann, Andreas; Ptacin, Jerod L; Grover, Ginni; Quirin, Sean; von Diezmann, Lexy; Lee, Marissa K; Backlund, Mikael P; Shapiro, Lucy et al. · Nano Lett · 2013

basic_science · Level V

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Abstract

We demonstrate quantitative multicolor three-dimensional (3D) subdiffraction imaging of the structural arrangement of fluorescent protein fusions in living Caulobacter crescentus bacteria. Given single-molecule localization precisions of 20-40 nm, a flexible locally weighted image registration algorithm is critical to accurately combine the super-resolution data with <10 nm error. Surface-relief dielectric phase masks implement a double-helix response at two wavelengths to distinguish two different fluorescent labels and to quantitatively and precisely localize them relative to each other in 3D.

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