Center for Microscopy, Microanalysis and Information Processing, University "Politehnica" Bucharest, 313 Splaiul Independentei, Sect. 6, Romania.
Microsc Res Tech. 2010 Mar;73(3):165-75. doi: 10.1002/jemt.20767.
Confocal laser scanning microscopy (CLSM) enables us to capture images representing optical sections on the volume of a specimen. The images acquired from different layers have a different contrast: the images obtained from the deeper layers of the specimen will have a lower contrast with respect to the images obtained from the topmost layers. The main reasons responsible for the effects described above are light absorption and scattering by the atoms and molecules contained in the volume through which the light passes. Also light attenuation can be caused by the inclination of the observed surface. In the case of the surfaces that have a steep inclination, the reflected light will have a different direction than the one of the detector. We propose a technique of digital image processing that can be used to compensate the effects of light attenuation based on histogram operations. We process the image series obtained by CLSM by exact histogram specification and equalization. In this case, a strict ordering among pixels must be induced in order to achieve the exact histogram modeling. The processed images will end up having exactly the specified histogram and not a histogram with a shape that just resembles to the specified one, as in the case of classical histogram specification algorithms. Experimental results and theoretical aspects of the induced ordering are discussed, as well as a comparison between several histogram modeling techniques with respect to the processing of image series obtained by confocal microscopy.
共聚焦激光扫描显微镜 (CLSM) 使我们能够捕捉代表样本体积的光学切片的图像。从不同层获得的图像具有不同的对比度：与从最顶层获得的图像相比，从样本更深层获得的图像对比度较低。导致上述影响的主要原因是光通过的体积中的原子和分子的光吸收和散射。此外，观察表面的倾斜也会导致光衰减。在具有陡峭倾斜度的表面的情况下，反射光的方向将与探测器的方向不同。我们提出了一种数字图像处理技术，可以用于基于直方图操作来补偿光衰减的影响。我们通过精确的直方图规定和均衡来处理 CLSM 获得的图像系列。在这种情况下，为了实现精确的直方图建模，必须在像素之间引入严格的排序。处理后的图像最终将具有完全指定的直方图，而不是具有指定形状的直方图，就像经典直方图规定算法的情况一样。讨论了所诱导排序的实验结果和理论方面，以及几种直方图建模技术在处理共焦显微镜获得的图像系列方面的比较。
