Department of Medical Technology and Clinical Engineering, Faculty of Health and Medical Sciences, Hokuriku University, Kanazawa, Japan.
Department of Medical Technology, Faculty of Health Sciences, Ehime Prefectural University of Health Sciences, Tobe, Japan.
Acta Cytol. 2021;65(6):510-521. doi: 10.1159/000518452. Epub 2021 Sep 14.
INTRODUCTION/OBJECTIVE: Liquid-based cytology (LBC) is advantageous as multiple stained specimens can be prepared and used for additional assays such as immunocytochemical and molecular-pathological investigations. Two types of preservative-fixative solutions (fixatives) are used for nongynecologic specimens used in the BD SurePath-LBC (SP-LBC) method, and their components vary. However, few studies have evaluated the differences in antigen-retaining ability between these fixatives. Therefore, we investigated and compared the antigen-retaining ability of the fixatives in immunocytochemical staining (ICC) under long-term storage conditions.
Sediments of cultured RAJI cells (derived from Burkitt's lymphoma) were added to each fixative (red and blue) and stored at room temperature for a specified period (1 h; 1 week; and 1, 3, and 6 months). The specimens were then prepared using the SP-LBC method and subjected to ICC. Positivity rate was calculated using the specimens fixed at room temperature for 1 h as a control. Antibodies against Ki67 expressed in the nucleus and against CD20 and leukocyte common antigen (LCA) expressed on the cell membrane were used.
For CD20 and LCA, the positivity rate increased with time in the red fixative compared with that in the control. In the blue fixative, the positivity rate was highest at 1 h and was maintained at a high level throughout the storage period. In contrast, the Ki67 positivity rate was highest at 1 h in both red and blue fixatives and markedly decreased with time. Therefore, although refrigerated (8°C) storage was used, no improvement was noted.
Long-term storage is possible for cell membrane antigens at room temperature; however, it is unsuitable for intranuclear antigens. Therefore, we conclude that suitable fixative type and storage temperature differ based on antigen location. Further investigation is warranted.
简介/目的：液基细胞学（LBC）具有优势，因为可以制备和使用多个染色标本进行额外的检测，如免疫细胞化学和分子病理学研究。BD SurePath-LBC（SP-LBC）方法中使用了两种类型的保存固定液（固定剂）来固定非妇科标本，它们的成分有所不同。然而，很少有研究评估这些固定剂在保持抗原方面的差异。因此，我们研究并比较了这两种固定剂在长期储存条件下进行免疫细胞化学染色（ICC）时的抗原保留能力。
将培养的 RAJI 细胞（源自伯基特淋巴瘤）的沉淀物添加到每种固定剂（红色和蓝色）中，并在室温下储存指定的时间（1 小时、1 周、1、3 和 6 个月）。然后使用 SP-LBC 方法制备标本，并进行 ICC。以室温固定 1 小时的标本作为对照计算阳性率。使用核内表达 Ki67 和细胞膜上表达的 CD20 和白细胞共同抗原（LCA）的抗体。
对于 CD20 和 LCA，与对照相比，红色固定剂中的阳性率随时间的推移而增加。在蓝色固定剂中，1 小时时的阳性率最高，并在整个储存期间保持较高水平。相比之下，红色和蓝色固定剂中 Ki67 的阳性率在 1 小时时最高，随着时间的推移显著降低。因此，尽管使用了冷藏（8°C）储存，但没有改善。
室温下可以长期储存细胞膜抗原，但不适合核内抗原。因此，我们得出结论，合适的固定剂类型和储存温度取决于抗原的位置。需要进一步研究。
