Design of a Scanning Module in a Confocal Microscopic Imaging System for Live-Cell Imaging

Abstract

This study proposes a Nipkow-based pinhole disk laser scanning confocal microscopic imaging system for ordinary optical microscopy, fluorescence microscopy, and confocal microscopy imaging of biological samples in order to realize the dynamic experimental monitoring of space-based life science experiments and the fine observation of biological samples. Confocal microscopic imaging is mainly completed by a scanning module that is composed of a spinning disk and other components. The parameters of the spinning disk directly determine the quality of the image. During the design process, the resolution and signal-to-noise ratios caused by different pinhole diameters in the spinning disk are the main considerations. Changes and image blurring caused by crosstalk due to the pinhole arrangement and different pinhole spacings are addressed. The high photon efficiency of the new EMCCD (electron-multiplying charge-coupled device) and CMOS (complementary metal-oxide-semiconductor) camera reduces the exposure time as much as possible, reduces damage to living cells, and achieves high-speed confocal imaging. It is shown in a confocal imaging experiment with a variable magnification of 1–40× that the imaging resolution of the system can reach a maximum of 2592 × 1944, the spatial resolution can reach 1 μm, and the highest sampling frequency is 10 fps, thus meeting the design requirements for high-speed live-cell imaging.