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Abstract：
The choice of fixation method significantly impacts tissue morphology and protein visualization after immunohistochemistry (IHC). In this study, we compared the effects of paraformaldehyde (PFA) and trichloroacetic acid (TCA) fixation prior to IHC on chicken embryos. Our findings underscore the importance of validating fixation methods for accurate interpretation of IHC results, with implications for antibody validation and tissue-specific protein localization studies. We found that TCA fixation resulted in larger and more circular nuclei compared to PFA fixation. Additionally, TCA fixation altered the appearance of subcellular localization and fluorescence intensity of various proteins, including transcription factors and cytoskeletal proteins. Notably, TCA fixation revealed protein localization domains that may be inaccessible with PFA fixation. These results highlight the need for optimization of fixation protocols depending on the target epitope and model system, emphasizing the importance of methodological considerations in biological analyses.
摘要：
固定方法的选择显着影响免疫组织化学（IHC）后的组织形态和蛋白质可视化。在这项研究中,我们比较了IHC前多聚甲醛（PFA）和三氯乙酸（TCA）固定对鸡胚的影响。我们的发现强调了验证固定方法对准确解释IHC结果的重要性，对抗体验证和组织特异性蛋白质定位研究具有重要意义。我们发现，与PFA固定相比，TCA固定导致更大，更圆形的核。此外,TCA固定改变了各种蛋白质的亚细胞定位和荧光强度的外观，包括转录因子和细胞骨架蛋白。值得注意的是,TCA固定显示PFA固定可能无法进入的蛋白质定位域。这些结果突出了根据靶表位和模型系统优化固定方案的需要。强调生物学分析中方法学考虑的重要性。
