Abstract
We demonstrate a novel approach for the real time visualization and quantification of the 3D spatial frequencies in an image domain. Our approach is based on the spectral encoding of spatial frequency principle and permits the formation of an image as a color map in which spatially separated spectral wavelengths correspond to the dominant 3D spatial frequencies of the object. We demonstrate that our approach can visualize and analyze the dominant axial internal structure for each image point in real time and with nanoscale sensitivity to structural changes. Computer modeling and experimental results of instantaneous color visualization and quantification of 3D structures of a model system and biological samples are presented.
Highlights
Microscopic imaging and accurate nanoscale quantification of the internal structures of an unlabeled three-dimensional (3D) object are of great importance to many fields, such as biology, medicine, materials science, and nanotechnology
The dominant colors in each Rt-spectral encoding of spatial frequency (SESF) image and in the corresponding Fourier plane are directly correlated with the dominant spatial frequency of the axial structure, which depends on the nanosphere size
We introduce a novel approach, spectral encoding of spatial frequency (Rt-SESF) for realtime imaging and quantitative characterization of the internal structure of a 3D object with nanoscale sensitivity
Summary
Microscopic imaging and accurate nanoscale quantification of the internal structures of an unlabeled three-dimensional (3D) object are of great importance to many fields, such as biology, medicine, materials science, and nanotechnology. Other methods to quantify structural information without reconstruction of the scattering potential include imaging light-scattering spectroscopy that utilizes spectral or angular elastic scattering dependences [11,12,13,14,15,16,17,18] and direct analysis of the Fourier spectrum of the object [19, 20]; methods based on the direct analysis of the Fourier spectrum [20] have been reported to provide less than 10 nm sensitivity to structural changes of 2D periodic objects Despite these advances, current techniques do not support real-time probing of 3D structures, especially at the nanoscale, due to limited resolution, accuracy, and sensitivity, especially in the axial direction. A wide bandwidth of spatial frequencies can be simultaneously passed through a low-NA optical system, allowing the observation of finer details that may not be resolved using conventional microscopy Based on this principle, we recently developed a new technique, spectral contrast imaging microscopy (SCIM), for real-time, super-resolution imaging [28]. We provide a brief theoretical presentation of our approach and demonstrate using model systems and biological samples the ability of SESF to achieve real-time quantitative structural imaging and characterization of 3D objects
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