Jacobs N, Giannini S L, Al-Saleh W, Hubert P, Boniver J, Delvenne P
University of Liège, Department of Pathology, Belgium.
J Immunol Methods. 1999 Feb 1;223(1):123-9. doi: 10.1016/s0022-1759(98)00205-1.
In this study, we have developed a simple and efficient technique for the isolation of viable lymphocytes from the epithelium and stroma of small pre-neoplastic squamous intraepithelial lesions (SIL) of the uterine cervix. Following the separation of the epithelium from the stroma using dispase II, both biopsy fragments were used to generate T lymphocytes. The stroma-derived lymphocytes were obtained by collecting and culturing the cells migrating out of the biopsy in the presence of IL2 (50 U/ml). An average of 0.7 x 10(6) and 1.4 x 10(6) lymphocytes could be obtained after 20 and 30 days of culture, respectively. For the expansion of lymphocytes derived from the pre-neoplastic epithelium (SIL) it was necessary to use a combination of irradiated peripheral blood mononuclear cells (PBMC) as a feeder layer with PHA (0.1%), in addition to IL2 (50 U/ml). Interestingly, these lymphocytes could be obtained using either allogeneic or syngeneic PBMCs. With this protocol, we were able to generate up to 100 x 10(6) lymphocytes from the epithelium, the majority of which were T lymphocytes.
在本研究中，我们开发了一种简单有效的技术，用于从子宫颈小的癌前鳞状上皮内病变（SIL）的上皮和基质中分离活淋巴细胞。使用 Dispase II 将上皮与基质分离后，两个活检碎片都用于生成 T 淋巴细胞。通过收集和培养在 IL2（50 U/ml）存在下从活检中迁移出来的细胞，获得基质来源的淋巴细胞。培养 20 天和 30 天后，平均分别可获得 0.7×10⁶ 和 1.4×10⁶ 个淋巴细胞。为了扩增癌前上皮（SIL）来源的淋巴细胞，除了 IL2（50 U/ml）外，还需要使用经辐照的外周血单个核细胞（PBMC）作为饲养层与 PHA（0.1%）组合。有趣的是，使用同种异体或同基因 PBMC 均可获得这些淋巴细胞。通过该方案，我们能够从上皮中生成多达 100×10⁶ 个淋巴细胞，其中大多数是 T 淋巴细胞。
