Optimizing microlens arrays for incoherent HiLo microscopy

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HiLo microscopy is a powerful, low-cost, and easily configurable technique for acquiring high-contrast optically-sectioned images. However, traditional HiLo microscopes are based on coherent light sources with diffusive glass plates or incoherent light sources with digital mirror devices (DMD) and spatial light modulators (SLM), which are more expensive. Here, we propose a new low-cost HiLo microscopy technique using MLAs and incoherent LED light sources. We simulated structured illumination (SI) patterns and HiLo image generation. To observe how MLAs affect HiLo images, we used three common MLAs with specific microlens pitch and numerical aperture (NA) to generate periodic illumination patterns. Our simulations concluded that the MLA's NA does not significantly affect HiLo images, and a larger lens pitch can bring a higher image contrast. However, there is an optimized lens pitch. If the lens pitch is equal to or higher than 140μm, artifacts are observed in HiLo images. Based on our simulations, employing the cross-type MLA with a 0.01 NA and a period of 120μm yields the optimal HiLo image (94.6% contrast). In our quantitative analysis with the traditional HiLo, where SI patterns were set with the same period, we calculated the SSIM index between each HiLo-Ground Truth image pair. The cross-type MLA HiLo performed slightly better (SSIM=0.9554) than the traditional HiLo (SSIM=0.9396). However, when using the hexagon-type MLA, artifacts were evident in the final HiLo image (SSIM=0.9054). The cylinder-type MLA HiLo result (SSIM=0.9366) is similar to the traditional HiLo. To our knowledge, this is the first numerical study about MLA-based HiLo microscopy. This study can benefit researchers using MLAs and incoherent light sources to configure their low-cost HiLo microscopes.
Original languageEnglish
Article number100622
Number of pages20
JournalResults in Optics
Early online date16 Feb 2024
Publication statusE-pub ahead of print - 16 Feb 2024


  • HiLo microscopy
  • simulation
  • incoherent fluorescent imaging
  • optically-sectioning


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