Focusing method and device for fluorescence microscope system

Abstract

The invention discloses a focusing method and device for a fluorescence microscope system. The method comprises the following steps that in the focus region selection step, a plurality of clear-focusfeature regions are selected during axial scanning of a fluorescent sample, and clear images and positions of each feature region are recorded; in the angle-axial position iterative adjustment optimization step, the movement amount of the angle and the axial position is obtained according to the central coordinate of each focus area, image matching alignment is performed after moving, the center coordinate of each focus area is measured, and then iterative optimization is conducted again to achieve focus. The method is suitable for a case that the sample changes during the adjustment process,is suitable for wide field fluorescence imaging, and effectively improves the focusing speed and stability.

Description

technical field [0001] The invention relates to the technical field of microscope system focusing, in particular to a focusing method and device for a fluorescence microscope system. Background technique [0002] Fluorescence microscopic imaging has important applications in the field of biomedical research, especially in the fields of clinical diagnosis and other fields, which has been paid more and more attention by people. Fluorescence microscopy often uses a flat detector to image the sample, and only the focal plane of the objective lens can be clearly imaged on the detector. Therefore, it is necessary to develop focusing techniques to make the target plane of the sample coincide with the focal plane of the microscope. [0003] The current automatic microfocus technology can be roughly divided into active focus technology and passive focus technology. The former actively emits infrared or ultrasonic to detect the position of the target object, and mechanically adjusts...

AI Technical Summary

Problems solved by technology

[0004] The active autofocus method cannot be directly used in fluorescence microscopy: due to the characteristics of fluorescence, the surface detected by infrared or ultrasound cannot be guaranteed to coincide with the fluorescent target plane that needs to be focused

Existing passive autofocus methods applied to fluorescence microscopy have three important defects: 1. The sharpness function of the image depends on the unimodal nature of the probe sample, that is, the objective function reaches its maximum value only on the target plane

This is often difficult to meet in fluorescence microscopy samples with three-dimensional structures

2. Most of the existing detection technologies use the stepping method to detect and find the clearest position of the image

In summary, the existing automatic microfocus technology has defects in fluorescence microscopy

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