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arXiv · 2511.20890

An Affordable Fiducial Marker Strategy for Reliable Autofocus in Long-Term Live Microscopy

Ilyas Djafer-Cherif·Bartlomiej Waclaw
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Abstract

Long-term time-lapse imaging of biological samples requires correcting for focal drift, which would otherwise gradually push the sample out of focus. We present a software-based method that eliminates this time-dependent blur using only a motorized Z-drive, with no additional hardware. The method relies on imaging marks made on the side of the coverslip opposite to the sample. We provide a Beanshell script implementation, evaluate its performance across multiple objectives, and benchmark it against a hardware autofocus system, finding comparable results. Finally, we demonstrate its effectiveness in live imaging of growing bacterial colonies.

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Ilyas Djafer-Cherif, Bartlomiej Waclaw. 2025-11-25. An Affordable Fiducial Marker Strategy for Reliable Autofocus in Long-Term Live Microscopy. https://doi.org/10.1111/jmi.70127

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