Structured Illumination for Super Resolution Imaging

Structured illumination microscopy is a technique based on the ability to get the high frequency information which is not available normally with optical system which has low bandwidths then we just have to transfer high frequency information into low bandwidth optics. In this technique, instead of illuminating a sample with the uniform field, the sample labelled by fluorescent material is illuminated with a structured pattern of high spatial frequency, generated by laser light passing through a movable optical grating and through objective lens projected onto the specimen, this result in the formation of coarse interference - moire fringes- in the Emission distribution, which encodes the high-frequency components into the "observable region" of the limited bandwidth of microscopic objective. Since one pattern is a purposely structured excitation light intensity whose frequency content is known, thus on illuminating the specimen in different orientation and processing all images acquired, it is possible using computer algorithms to compute the unknown frequency content of the sample. Thus, one can gain access to normally high-resolution information about the sample. This methodology requires computers to reconstruct the image and extract all the information present as in this microscopy image cannot be interpreted visually, as it is some type of coded image.

Although SR-SIM, in its original implementation, is limited to a factor of approximately 2 in improving the spatial resolution and unable to resolve transparent biological tissues or cells which is very important from medical perspective, this barrier can also be superseded with somewhat more sophisticated optical schemes.

For the study of structure and dynamics of sub-cellular components, there is a modification on SIM which offers higher resolution with extended field of view. In conventional SIM same objective lens is used for both illumination and recording purpose; on the other hand, if illumination technique is decoupled from imaging then it removes the limitation imposed by the aperture of lens and can achieve higher than 2-fold resolution. This is done by a new technique based on Transmission type SIM. To achieve high resolution information multiple steps are performed in chronological order. The beams from Spatial Light Modulator (SLM) are made to reflect from a set of mirrors, results in the formation of Interference pattern. This illumination pattern is used to illuminate the biological sample to be imaged. By changing the tilt angle of the set of mirrors, the frequency of the illumination pattern is changed, thus very high frequency information can be moved towards the pass band, collected through a diffraction limited objective. As the images recorded are in the coded form, by proper decoding algorithm, images processed computationally to go beyond the two-fold resolution of a reflection type conventional SIM.

Publications

Samanta, Krishnendu, et al. "Improving resolution in two orthogonal orientations from a single-shot digital holographic microscopy." Results in Optics 14 (2024): 100586.

Samanta, Krishnendu, et al. "Transmission structured illumination microscopy with tunable frequency illumination using tilt mirror assembly." Scientific Reports 13.1 (2023): 1453.

Samanta, Krishnendu, et al. "Saturable absorption assisted nonlinear structured illumination microscopy." Optics Letters 47.11 (2022): 2702-2705.

Samanta, Krishnendu, and Joby Joseph. "Image reconstruction approach for a high space-bandwidth product structured illumination microscopy system." JOSA A 38.12 (2021): 1744-1751.

Samanta, Krishnendu, and Joby Joseph. "An overview of structured illumination microscopy: recent advances and perspectives." Journal of Optics 23.12 (2021): 123002.

Samanta, Krishnendu, et al. "Blind super-resolution approach for exploiting illumination variety in optical-lattice illumination microscopy." ACS Photonics 8.9 (2021): 2626-2634.

Joseph, Joby, et al. "Improving the space-bandwidth product of structured illumination microscopy using a transillumination configuration." Journal of Physics D: Applied Physics 53.4 (2019): 044006.

Joseph, J., Faiz, K. P., Lahrberg, M., Tinguely, J. C., & Ahluwalia, B. S. (2019). Improving the space-bandwidth product of structured illumination microscopy using a ransillumination configuration. Journal of Physics D: Applied Physics, 53(4), 044006.