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The structure of the cell nucleus of higher organisms has become a major topic of advanced light microscopy. So far, a variety of methods have been applied, including confocal laser scanning fluorescence microscopy, 4Pi, STED and localisation microscopy approaches, as well as different types of patterned illumination microscopy, modulated either laterally (in the object plane) or axially (along the optical axis). Based on our experience, we discuss here some application perspectives of Modulated Illumination Microscopy (MIM) and its combination with single-molecule localisation microscopy (SMLM). For example, spatially modulated illumination microscopy/SMI (illumination modulation along the optical axis) has been used to determine the axial extension (size) of small, optically isolated fluorescent objects between ≤ 200 nm and ≥ 40 nm diameter with a precision down to the few nm range; it also allows the axial positioning of such structures down to the 1 nm scale; combined with laterally structured illumination/SIM, a 3D localisation precision of ≤1 nm is expected using fluorescence yields typical for SMLM applications. Together with the nanosizing capability of SMI, this can be used to analyse macromolecular nuclear complexes with a resolution approaching that of cryoelectron microscopy., (© 2024 The Author(s). Journal of Microscopy published by John Wiley & Sons Ltd on behalf of Royal Microscopical Society.)

Objectives: To assess the impact of posterior corneal asphericity on postoperative calculation error using the Haigis-L and the Barrett formulas for eyes after laser in situ keratomileusis or photorefractive keratectomy (PRK)., Methods: We assessed the mean absolute error (MAE) of two power calculation formulas, Barrett true-K and Haigis-L formulas, in a retrospective analysis of 34 eyes of 34 patients who underwent cataract surgery. We performed a regression analysis between corneal parameters (anterior and posterior Q values, Kmax, K1, and K2) and the MAE of each formula., Results: In the cohort, 11 eyes were of women and 23 of men. The average age of the study population was 66.5±8.6 years. The mean axial length was 24±4.7 mm, the mean anterior chamber depth was 3.27±0.7 mm, and the mean posterior Q-value was -0.15±0.28. The MAE of Haigis-L and Barrett true-K formulas were 0.72 and 0.68, respectively (P=0.54). The regression analysis showed a statistically significant relationship only between the error in refraction prediction and the posterior Q-value regardless of the formula used. The coefficient of determination was higher for the Barrett true-K formula (r=0.52; R2=0.28; P<0.05), compared with the Haigis-L (r=0.49; R2=0.25; P<0.05)., Conclusions: Posterior corneal surface asphericity influences the refractive error of calculation using both Haigis-L and Barrett true-K formulas for eyes after a myopic PRK or laser-assisted in situ keratomileusis surgery., Competing Interests: The authors have no funding or conflicts of interest to disclose., (Copyright © 2021 Contact Lens Association of Ophthalmologists.)

Thousands of genes and the complex biochemical networks for their transcription are packed in the micrometer sized cell nucleus. To control biochemical processes, spatial organization plays a key role. Hence the structure of the cell nucleus of higher organisms has emerged as a main topic of advanced light microscopy. So far, a variety of methods have been applied for this, including confocal laser scanning fluorescence microscopy, 4Pi-, STED- and localization microscopy approaches, as well as (laterally) structured illumination microscopy (SIM). Here, we summarize the state of the art and discuss application perspectives for nuclear nanostructure analysis of spatially modulated illumination (SMI). SMI is a widefield-based approach to using axially structured illumination patterns to determine the axial extension (size) of small, optically isolated fluorescent objects between less than or equal to 200 nm and greater than or equal to 40 nm diameter with a precision down to the few nm range; in addition, it allows the axial positioning of such structures down to the 1 nm scale. Combined with SIM, a three-dimensional localization precision of less than or equal to 1 nm is expected to become feasible using fluorescence yields typical for single molecule localization microscopy applications. Together with its nanosizing capability, this may eventually be used to analyse macromolecular complexes and other nanostructures with a topological resolution, further narrowing the gap to Cryoelectron microscopy. This article is part of the Theo Murphy meeting issue 'Super-resolution structured illumination microscopy (part 2)'.