Miller Lisa M, Wang Qi, Smith Randy J, Zhong Hui, Elliott Donald, Warren John
National Synchrotron Light Source, Brookhaven National Laboratory, Bldg 725 D, 75 Brookhaven Avenue, Upton, NY 11973, USA.
Anal Bioanal Chem. 2007 Mar;387(5):1705-15. doi: 10.1007/s00216-006-0879-2. Epub 2006 Nov 18.
Many disease processes involve alterations in the chemical makeup of tissue. Synchrotron-based infrared (IR) and X-ray fluorescence (XRF) microscopes are becoming increasingly popular tools for imaging the organic and trace metal compositions of biological materials, respectively, without the need for extrinsic labels or stains. Fourier transform infrared microspectroscopy (FTIRM) provides chemical information on the organic components of a material at a diffraction-limited spatial resolution of 2-10 microm in the mid-infrared region. The synchrotron X-ray fluorescence (SXRF) microprobe is a complementary technique used to probe trace element content in the same systems with a similar spatial resolution. However to be most beneficial, it is important to combine the results from both imaging techniques on a single sample, which requires precise overlap of the IR and X-ray images. In this work, we have developed a sample substrate containing a gold grid pattern on its surface, which can be imaged with both the IR and X-ray microscopes. The substrate consists of a low trace element glass slide that has a gold grid patterned on its surface, where the major and minor parts of the grid contain 25 and 12 nm gold, respectively. This grid pattern can be imaged with the IR microscope because the reflectivity of gold differs as a function of thickness. The pattern can also be imaged with the SXRF microprobe because the Au fluorescence intensity changes with gold thickness. The tissue sample is placed on top of the patterned substrate. The grid pattern's IR reflectivity image and the gold SXRF image are used as fiducial markers for spatially overlapping the IR and SXRF images from the tissue. Results show that IR and X-ray images can be correlated precisely, with a spatial resolution of less than one pixel (i.e., 2-3 microns). The development of this new tool will be presented along with applications to paraffin-embedded metalloprotein crystals, Alzheimer's disease, and hair composition.
许多疾病过程都涉及组织化学组成的改变。基于同步加速器的红外（IR）显微镜和X射线荧光（XRF）显微镜正日益成为分别对生物材料的有机成分和痕量金属成分进行成像的常用工具，无需使用外在标记或染色剂。傅里叶变换红外显微光谱（FTIRM）在中红外区域以2 - 10微米的衍射极限空间分辨率提供材料有机成分的化学信息。同步加速器X射线荧光（SXRF）微探针是一种互补技术，用于在相同系统中以类似的空间分辨率探测痕量元素含量。然而，为了获得最大益处，将两种成像技术对单个样本的结果相结合很重要，这需要红外图像和X射线图像精确重叠。在这项工作中，我们开发了一种表面带有金网格图案的样本基底，它可以用红外显微镜和X射线显微镜成像。该基底由一块低痕量元素载玻片组成，其表面有金网格图案，网格的主要部分和次要部分分别含有25纳米和12纳米的金。这种网格图案可以用红外显微镜成像，因为金的反射率随厚度变化。该图案也可以用SXRF微探针成像，因为金的荧光强度随金厚度变化。将组织样本放置在有图案的基底上。网格图案的红外反射率图像和金的SXRF图像用作基准标记，用于在空间上重叠来自组织的红外图像和SXRF图像。结果表明，红外图像和X射线图像可以精确关联，空间分辨率小于一个像素（即2 - 3微米）。将展示这种新工具的开发以及在石蜡包埋的金属蛋白晶体、阿尔茨海默病和毛发成分方面的应用。
