Schwertner M, Booth M, Wilson T
Opt Express. 2004 Dec 27;12(26):6540-52. doi: 10.1364/opex.12.006540.
Aberrations are known to severely compromise image quality in optical microscopy, especially when high numerical aperture (NA) lenses are used in confocal fluorescence microscopy (CFM) and two-photon microscopy (TPM). The method of adaptive optics may correct aberrations and restore diffraction limited operation. So far the problem of aberrations that occur in the imaging of biological specimens has not been quantified. However, this information is essential for the design of adaptive optics systems. We have therefore built an interferometer incorporating high NA objective lenses to measure the aberrations introduced by biological specimens. The measured wavefronts were decomposed into their Zernike mode content in order both to classify and quantify the aberrations. We calculated the potential benefit of correcting different numbers of Zernike modes using different NAs in an adaptive CFM by comparing the signal levels before and after correction. The results indicate that adaptive correction of low order Zernike modes can provide significant benefit for many specimens. The results also show that quantitative fluorescence microscopy may be strongly affected by specimen induced aberrations in non-adaptive systems.
众所周知，像差会严重影响光学显微镜的图像质量，尤其是在共聚焦荧光显微镜（CFM）和双光子显微镜（TPM）中使用高数值孔径（NA）透镜时。自适应光学方法可以校正像差并恢复衍射极限操作。到目前为止，生物样本成像中出现的像差问题尚未得到量化。然而，这些信息对于自适应光学系统的设计至关重要。因此，我们构建了一台包含高NA物镜的干涉仪，以测量生物样本引入的像差。将测量的波前分解为其泽尼克模式成分，以便对像差进行分类和量化。通过比较校正前后的信号水平，我们计算了在自适应CFM中使用不同NA校正不同数量泽尼克模式的潜在益处。结果表明，对低阶泽尼克模式进行自适应校正可为许多样本带来显著益处。结果还表明，在非自适应系统中，定量荧光显微镜可能会受到样本诱导像差的强烈影响。
