CAJ | 학술논문

目的：比较Percoll非连续密度梯度离心法、Ficoll-Hypaque 密度梯度离心法分离结直肠癌中的巨噬细胞的效果，并进一步探讨贴壁分离巨噬细胞的最适宜贴壁时间。方法：取结直肠癌患者的肿瘤组织，制成单个细胞的悬液，分别使用100％ Ficoll、35％ Percoll 以及25％与65％双梯度 Percoll 进行巨噬细胞分离，37℃培养2 h后，去除未贴壁的细胞，使用免疫荧光实验来检测巨噬细胞的纯度。之后，将Ficoll分离得到的单个核细胞，分别培养1、2、4和9 h后比较巨噬细胞的纯度。结果：Ficoll-Hypaque 密度梯度离心法分离的巨噬细胞的纯度为60．18％，Percoll 非连续密度梯度离心法分离的巨噬细胞的纯度分别为54．33％和10．93％，均低于Ficoll法分离所得到的巨噬细胞的纯度（P ＜0．05）。 Ficoll分离后贴壁1、2、4 h 分离得到的巨噬细胞纯度均达到60％以上，高于贴壁培养9 h得到的巨噬细胞的纯度（34．67％）。结论： Ficoll-Hypaque密度梯度离心法分离巨噬细胞，纯化程度好，是一种简单、高效的巨噬细胞分离方法，适于临床和科研中广泛应用。贴壁培养2～4h是贴壁分离巨噬细胞最适宜的时间。
목적：비교Percoll비련속밀도제도리심법、Ficoll-Hypaque 밀도제도리심법분리결직장암중적거서세포적효과，병진일보탐토첩벽분리거서세포적최괄의첩벽시간。방법：취결직장암환자적종류조직，제성단개세포적현액，분별사용100％ Ficoll、35％ Percoll 이급25％여65％쌍제도 Percoll 진행거서세포분리，37℃배양2 h후，거제미첩벽적세포，사용면역형광실험래검측거서세포적순도。지후，장Ficoll분리득도적단개핵세포，분별배양1、2、4화9 h후비교거서세포적순도。결과：Ficoll-Hypaque 밀도제도리심법분리적거서세포적순도위60．18％，Percoll 비련속밀도제도리심법분리적거서세포적순도분별위54．33％화10．93％，균저우Ficoll법분리소득도적거서세포적순도（P ＜0．05）。 Ficoll분리후첩벽1、2、4 h 분리득도적거서세포순도균체도60％이상，고우첩벽배양9 h득도적거서세포적순도（34．67％）。결론： Ficoll-Hypaque밀도제도리심법분리거서세포，순화정도호，시일충간단、고효적거서세포분리방법，괄우림상화과연중엄범응용。첩벽배양2～4h시첩벽분리거서세포최괄의적시간。
Objective Percoll density gradient centrifugation and Ficoll-Hypaque density gradient cen-trifugation, which are frequently-used methods for separation of tumor-associated macrophages (TAMs) from solid carcinoma were compared, in order to find an effective way to separate TAMs from colorectal carcinoma (CRC). Furthermore, we studied the best adherence time of separating macrophage among mononuclear cells. Methods specimens were collected from CRC patients , after digesting into single cells , TAMs were separated from the same specimen by 100% Ficoll, 35% percoll and 25% combined with 65% percoll respectively. After these pre-liminary separation, the collected cells were purified a second time by adherence separation. The purity of TAMs were detected by immunofluorescence. Results TAMs purity from Ficoll-Hypaque density gradient centrifugation was 80.18%, statistically higher than that from Percoll density gradient centrifugations (54.33% and 10.93% re-spectively). Conclusion Compared to Percoll density gradient centrifugation, Ficoll-Hypaque density gradient centrifugation is a more effective and simple way to isolate TAMs from colorectal carcinoma , suggesting it can be wildly used in clinical and basic medical research. 2-4 hours is the best adherence time for isolating macrophage.