Collagen has a major role in the structural organization of tendons. Picrosirius red (PSR) staining viewed under polarized light microscopy is the standard method to evaluate the organization of collagen fibers in tissues. It is also used to distinguish between type I and type III collagen in tissue sections. However, accurate analysis and interpretation of PSR images are challenging because of technical factors and historical misconceptions. The aim of this study was to clarify whether collagen types I and III can be distinguished by PSR staining in rat Achilles tendons, using double immunohistochemistry as the positive control. Our findings showed that PSR staining viewed with polarized light microscopy was suitable for qualitative and quantitative assessment of total collagen but was not able to distinguish collagen types. We found it critical to use a polarizing microscope equipped with a rotating stage; tendon section orientation at 45° with respect to crossed polarizers was optimal for the qualitative and quantitative assessment of collagen organization. Immunohistochemistry was superior to PSR staining for detection of collagen type III. We also compared formalin and Bouin solution as fixatives. Both produced similar birefringence, but formalin-fixed tendons provided higher quality histological detail with both hematoxylin–eosin and immunostaining:

胶原蛋白在肌腱的结构组织中起主要作用。在偏光显微镜下观察到的天狼星红 (PSR) 染色是评估组织中胶原纤维组织的标准方法。它还用于区分组织切片中的 I 型和 III 型胶原蛋白。然而，由于技术因素和历史误解，对 PSR 图像的准确分析和解释具有挑战性。本研究的目的是阐明是否可以通过 PSR 染色在大鼠跟腱中区分 I 型和 III 型胶原，使用双重免疫组织化学作为阳性对照。我们的研究结果表明，用偏光显微镜观察的 PSR 染色适用于总胶原蛋白的定性和定量评估，但不能区分胶原蛋白类型。我们发现使用配备旋转台的偏光显微镜至关重要。与交叉偏振器成 45° 的肌腱截面方向对于胶原组织的定性和定量评估是最佳的。对于 III 型胶原的检测，免疫组织化学优于 PSR 染色。我们还比较了福尔马林和布安溶液作为固定剂。两者都产生了相似的双折射，但福尔马林固定的肌腱通过苏木精-伊红和免疫染色提供了更高质量的组织学细节：我们还比较了福尔马林和布安溶液作为固定剂。两者都产生了相似的双折射，但福尔马林固定的肌腱通过苏木精-伊红和免疫染色提供了更高质量的组织学细节：我们还比较了福尔马林和布安溶液作为固定剂。两者都产生了相似的双折射，但福尔马林固定的肌腱通过苏木精-伊红和免疫染色提供了更高质量的组织学细节：