Advancecor GmbH, 82152, Martinsried, Germany.
Sci Rep. 2020 Nov 16;10(1):19849. doi: 10.1038/s41598-020-76950-1.
A novel Histo-ELISA technique is intended to facilitate quantification of target tissue proteins in a tissue section and involves the selection of target regions in the tissue section, application of streptavidin-conjugated HRP (horseradish peroxidase), coupled with peroxidase substrate-TMB (3,3',5,5'-tetramethylbenzidine), and staining dye evaluation with ELISA reader. The target protein content (weight per volume unit) was translated from optical densities by a reference standard curve, obtained via parallel staining of the targeted protein-coated slides. To validate the technique, we carried out quantifications of IgG extravasation in ischemic and nonischemic brain sections in a mouse stroke model. With those obtained data and the reference of immunohistochemistry scores assessed on the adjacent sections, accuracy, sensitivity, and precision for the technique were evaluated. For all evaluated parameters, Histo-ELISA performance was either comparable to or better than the standard immunohistochemistry. A comparison with the data from the repeated measurements yielded a rather low coefficient of variation. The results confirmed that the technique is a fairly reliable quantitative test with rather high sensitivity, accuracy, precision, and reproducibility for detecting target protein content in tissue sections and that its tissue distribution and related subsequent morphological changes can be observed at the same time.
一种新的组织酶联免疫吸附测定（Histo-ELISA）技术旨在促进组织切片中目标组织蛋白的定量，并涉及在组织切片中选择目标区域，应用链霉亲和素缀合的辣根过氧化物酶（HRP），与过氧化物酶底物-TMB（3,3',5,5'-四甲基联苯胺）偶联，并使用 ELISA 读取器评估染色染料。目标蛋白含量（体积单位重量）通过参考标准曲线从光密度值转换得出，该曲线通过靶向蛋白包被载玻片的平行染色获得。为了验证该技术，我们在小鼠中风模型中对缺血和非缺血脑组织切片中的 IgG 渗出进行了定量。利用获得的数据和相邻切片上评估的免疫组织化学评分，评估了该技术的准确性、敏感性和精密度。对于所有评估的参数，Histo-ELISA 的性能要么与标准免疫组织化学相当，要么优于标准免疫组织化学。与重复测量数据的比较产生了相当低的变异系数。结果证实，该技术是一种相当可靠的定量检测方法，具有相当高的敏感性、准确性、精密度和重现性，可用于检测组织切片中的目标蛋白含量，并且可以同时观察其组织分布和相关的后续形态变化。
