Faculdade São Leopoldo Mandic, São Leopoldo Mandic Institute and Research Center, Laboratory of Neuroimmune Interface of Pain Research, Rua José Rocha Junqueira, 13 - Swift - Zip Code 13045-755, Campinas, São Paulo, Brazil.
Faculdade São Leopoldo Mandic, São Leopoldo Mandic Institute and Research Center, Laboratory of Pathology, Rua José Rocha Junqueira, 13 - Swift - Zip Code 13045-755, Campinas, São Paulo, Brazil.
Arch Oral Biol. 2021 Jun;126:105115. doi: 10.1016/j.archoralbio.2021.105115. Epub 2021 Mar 30.
Propose a standard, fast and accurate protocol for the processing of the temporomandibular joint (TMJ) of adults' rats for histology and immunohistochemistry reactions.
Wistar male rats were perfused with paraformaldehyde (4 %). The heads were fixed in formaldehyde 10 % solution for 48 h. After that, the heads were sectioned in a sagittal plane and fixed for plus 48 h. Decalcification was performed using 20 % formic acid for 96 h and delimitation of TMJ area was done. Detailed methodology to a standard extraction and processing of TMJ to histological sections is described. Different buffers, equipment, temperature and time were tested to optimize immunostaining. Morphological preservation and antigenicity were evaluated by hematoxylin and eosin staining and immunohistochemistry reaction.
The current findings demonstrated that TMJ fixed in 10 % formaldehyde and decalcified in 20 % formic acid optimized decalcification processing time with preservation of cell morphology. Antigen retrieval with citrate buffer in pressure cooker (2 min at 100 °C and 5 min at room temperature) demonstrated the best protocol to preservation of the structures of TMJ.
This work demonstrates in detail a methodology of a fast and accurate TMJ processing for histology and immunohistochemistry reactions that guarantee tissue integrity and quality of staining.
提出一种标准、快速、准确的处理成年大鼠颞下颌关节（TMJ）进行组织学和免疫组织化学反应的方案。
雄性 Wistar 大鼠用多聚甲醛（4%）灌注。头部用 10%甲醛溶液固定 48 小时。然后，头部在矢状面切片并固定 48 小时。使用 20%甲酸进行 96 小时脱钙，并对 TMJ 区域进行界定。详细描述了一种标准的 TMJ 提取和处理方法，以获得组织学切片。测试了不同的缓冲液、设备、温度和时间，以优化免疫染色。通过苏木精和伊红染色和免疫组织化学反应评估形态保存和抗原性。
目前的研究结果表明，用 10%甲醛固定和用 20%甲酸脱钙可以优化脱钙处理时间，同时保持细胞形态。在高压锅（100°C 2 分钟和室温 5 分钟）中用柠檬酸盐缓冲液进行抗原修复显示了保存 TMJ 结构的最佳方案。
本工作详细介绍了一种快速、准确的 TMJ 处理方法，用于组织学和免疫组织化学反应，保证了组织的完整性和染色质量。
