基于点扫描结构光的SHG超分辨显微技术研究

SHG Super-Resolution Microscopy Based on Point-Scanning Structured Light

二次谐波产生(SHG)成像技术是一种针对非中心对称生物组织的免标记成像技术,已经成为生命科学研究的重要手段。衍射极限使得SHG技术无法分辨衍射极限以下的精细结构。虽然超分辨显微技术取得了突破性进展,但是SHG的相干非线性过程限制了SHG超分辨显微技术的发展。提出了一种点扫描结构光照明SHG超分辨显微(SHG-psSIM)技术,实现了氧化锌颗粒和小鼠尾腱的超分辨SHG显微成像。在传统的SHG显微系统的激发光路中引入电光调制器,通过对激发光正弦调制产生点扫描结构照明图案。基于点扫描结构照明图案与样本结构相互作用产生的莫尔条纹效应,将原本不可探测的样本高频信息搬移到显微镜通频带内,并利用光电倍增管探测。最后,利用软件重构出超分辨率图像。对比传统SHG系统,SHG-psSIM分辨率提高了1.86倍。

Second harmonic generation(SHG)imaging technology is a powerful tool capable of imaging non-centrosymmetric biological tissues without labeling,which has become an important means of life science research.Due to the diffraction limit,SHG technology cannot resolve the fine structures below the diffraction limit.Although super-resolution microscopy has made breakthrough progress,the coherent nonlinear process of SHG limits the development of the SHG super-resolution microscopy.A point-scanning structured illumination SHG super-resolution microscopy(SHG-psSIM)technology is proposed to realize the super-resolution SHG microscopic imaging of zinc oxide particles and mouse tail tendons.The electro-optic modulator is introduced into the excitation light path of the traditional SHG microscopy system.A point-scanning structured illumination pattern is generated by sinusoidally modulating of the excitation beam.Based on the moiréfringe effect generated by interaction of the point-scanning structured illumination pattern with the sample structures,the undetectable high frequency information of samples is moved to the observable passband of the microscopy and detected by the photomultiplier.Finally,the algorithm software is used to reconstruct the super-resolution image.Compared with the traditional SHG system,the resolution of SHG-psSIM is improved by 1.86 times.