Simple autofocusing method by image processing using transmission images for large-scale two-photon lithography.

We propose a simple autofocusing technique that can be introduced into conventional two-photon lithography systems without additional devices. Autofocusing is achieved by image processing using transmission images of photopolymerized voxels. The signal-to-noise ratio of transmission images was improved by optimal low-pass filtering to detect voxels in them. The focal point was detected with an accuracy of about 250 nm from the difference images. Further, we demonstrated mass-fabrication of a 5 × 5 spiral square array with an area of 900 × 900 µm2 using this method. The method has potential application in constructing low-cost, compact and versatile two-photon lithography apparatus.