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| Main Authors: | , , , , , , , , |
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| Format: | Preprint |
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2026
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| Subjects: | |
| Online Access: | https://arxiv.org/abs/2603.16908 |
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| _version_ | 1866912972034539520 |
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| author | Qian, Jiaming Feng, Jing Wu, Hongjun Zhang, Maoxian Lu, Dongqin Kang, Tianchi Han, Xinyu Chen, Qian Zuo, Chao |
| author_facet | Qian, Jiaming Feng, Jing Wu, Hongjun Zhang, Maoxian Lu, Dongqin Kang, Tianchi Han, Xinyu Chen, Qian Zuo, Chao |
| contents | Structured illumination microscopy (SIM) has emerged as a widely adopted super-resolution fluorescence imaging modality, offering high speed, low phototoxicity, large field-of-view, and compatibility with conventional probes. However, when applied to thick or scattering specimens, conventional two-dimensional SIM (2D-SIM) suffers from the missing cone problem in its optical transfer function, resulting in prominent out-of-focus background and severe reconstruction artifacts that compromise image fidelity. Here, we present four-beam interference structured illumination microscopy (4I-SIM), which introduces additional interference orders to expand lateral frequency support and compensate the axial missing cone simultaneously. This strategy achieves artifact-free super-resolution with intrinsic optical sectioning, effectively overcoming the fundamental limitation of 2D-SIM without additional acquisition overhead. Experimental validation across diverse thick fixed and live specimens demonstrates that 4I-SIM delivers nearly twofold lateral resolution enhancement and substantially improved sectioning compared with its 2D counterpart, achieving lateral and axial resolutions of 103 nm and 336 nm, respectively. In particular, 4I-SIM reveals mitochondrial remodeling and apoptosis under high-glucose stress with millisecond temporal resolution -- features that remain obscured with conventional SIM. With minimal hardware modification, low phototoxicity, and open-source reconstruction tools, 4I-SIM establishes a practical and reproducible platform for simultaneous super-resolution and optical sectioning imaging in complex biological environments. |
| format | Preprint |
| id |
arxiv_https___arxiv_org_abs_2603_16908 |
| institution | arXiv |
| publishDate | 2026 |
| record_format | arxiv |
| spellingShingle | Simultaneous super-resolution and optical sectioning with four-beam interference structured illumination microscopy (4I-SIM) Qian, Jiaming Feng, Jing Wu, Hongjun Zhang, Maoxian Lu, Dongqin Kang, Tianchi Han, Xinyu Chen, Qian Zuo, Chao Image and Video Processing Structured illumination microscopy (SIM) has emerged as a widely adopted super-resolution fluorescence imaging modality, offering high speed, low phototoxicity, large field-of-view, and compatibility with conventional probes. However, when applied to thick or scattering specimens, conventional two-dimensional SIM (2D-SIM) suffers from the missing cone problem in its optical transfer function, resulting in prominent out-of-focus background and severe reconstruction artifacts that compromise image fidelity. Here, we present four-beam interference structured illumination microscopy (4I-SIM), which introduces additional interference orders to expand lateral frequency support and compensate the axial missing cone simultaneously. This strategy achieves artifact-free super-resolution with intrinsic optical sectioning, effectively overcoming the fundamental limitation of 2D-SIM without additional acquisition overhead. Experimental validation across diverse thick fixed and live specimens demonstrates that 4I-SIM delivers nearly twofold lateral resolution enhancement and substantially improved sectioning compared with its 2D counterpart, achieving lateral and axial resolutions of 103 nm and 336 nm, respectively. In particular, 4I-SIM reveals mitochondrial remodeling and apoptosis under high-glucose stress with millisecond temporal resolution -- features that remain obscured with conventional SIM. With minimal hardware modification, low phototoxicity, and open-source reconstruction tools, 4I-SIM establishes a practical and reproducible platform for simultaneous super-resolution and optical sectioning imaging in complex biological environments. |
| title | Simultaneous super-resolution and optical sectioning with four-beam interference structured illumination microscopy (4I-SIM) |
| topic | Image and Video Processing |
| url | https://arxiv.org/abs/2603.16908 |