High-Resolution Oil-Free Imaging of Chromosomes Using Fourier Ptychographic Microscopy

Abstract

In conventional microscopic imaging of chromosomes, the use of high numerical aperture (NA) oil-immersion objectives is essential. However, the use of oil-immersion objectives poses a significant challenge to automated imaging systems because it increases the risk of sample contamination and instrumental damage. The shallow depth of field (DOF) of oil-immersion objectives also demands more sophisticated mechanical focusing. Here, we introduce a chromosome oil-free microscopic imaging system based on Fourier Ptychographic Microscopy (FPM) technology. The system employs a 100×, 0.8 NA dry objective to achieve a half-pitch resolution of 194 nm at an incident wavelength of 524 nm. The reconstructed images of chromosomes surpassed the conventional imaging with a 100×, 1.25 NA oil-immersion objective. We also utilized digital refocusing methods to extend the effective DOF to ±2.4 μm. This study preliminarily validates the possibility of developing a new generation of chromosome scanners without using an oil-immersion objective.