Harmonic microscopic measurement method based on sheet-light microscopy and confocal slit detection

Abstract

The invention discloses a harmonic microscopic measurement method based on sheet-light microscopy and confocal slit detection, and belongs to the field of non-linear optical measurement. In the harmonic microscopy measurement, the detection direction of a harmonic signal is perpendicular to the illumination direction of a sample, so that sheet-light measurement in harmonic microscopy is achieved.Confocal slit detection is achieved by using sCMOS as a detector and a rolling shutter working mode. A femtosecond laser pulse is reflected by a scanning galvanometer, then enters a switching opticalsystem for spherical aberration compensation, and is converged in the sample by a microscopic objective to form an excitation focusing light -spot needed by harmonic signal generation. The harmonic signal excited by the sample is collected by a detecting objective perpendicular to the illumination direction, and then passes through a spike filter to filter stray light, and is received for detection by the sCMOS working in a rolling shutter mode. The scanning process and rolling shutter docking are synchronic, and harmonic images with different position pixels are compounded by algorithms. According to the harmonic microscopic measurement method, the contrast ratio and the signal-to-noise ratio of harmonic microscopic imaging can be effectively increased, and harmonic microscopic imaging with the high frame rate is achieved.

Description

technical field [0001] The invention belongs to the field of optical microscopic measurement, and mainly relates to an ultra-precise non-contact measurement method for measuring three-dimensional fine structures in nano devices and biological samples. Background technique [0002] By using the nonlinear optical effects of the sample itself, such as second harmonic generation and third harmonic generation, microscopic imaging of biological samples without fluorescent labels, microstructure detection of nano-devices, and diagnosis of disease mechanisms can be performed. However, in harmonic microscopy imaging, due to the weak harmonic signal, it is often submerged in background noise, and optical scattering in thick samples, resulting in reduced contrast and resolution of harmonic imaging. In confocal scanning microscopy methods, pinholes can be used to eliminate out-of-focus stray light. However, this point-by-point scanning method reduces the image acquisition rate and lead...

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Problems solved by technology

However, this point-by-point scanning method reduces the image acquisition rate, resulting in long-term exposure of the sample, which is not conducive to microscopic imaging of living samples.

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