Chen W F, Scollay R, Shortman K
Thymus. 1983 Apr;5(3-4):197-207.
The frequency of all precursors of T cells capable of proliferation (PTL-p), and of all precursors of cytotoxic T-cell clones (CTL-p), was determined for mouse thymic and peripheral T-cell subsets differing in Ly phenotype. A high cloning efficiency, concanavalin A (Con A) and growth factor driven limit dilution culture system was used. A lectin-mediated non-specific lysis readout was used for detecting cytotoxic clones. This approach provided a balance sheet of the overall distribution of functional cells regardless of specificity. Subsets of splenic T lymphocytes were isolated by fluorescence-activated cell sorting (FACS) after two-colour staining with monoclonal anti-Thy 1 and anti-Ly 2 antibodies. Both the Ly 1+2- and Ly 1+2% subsets responded by clonal proliferation, but cytotoxic activity was almost exclusively limited to the Ly 1+2% derived clones. Four subclasses of thymocytes were isolated by FACS after two-colour staining with peanut agglutinin (PNA) and monoclonal anti-Ly 2 antibody. These were PNA+Ly 2+, PNA+ Ly 2-, PNA- Ly 2+ and PNA- Ly 2-, representing 80, 5, 5 and 10% of total thymocytes, respectively. Their respective PTL-p frequencies were 1 in 333, 1 in 200, 1 in 5.3 and 1 in 3.2, values which included a significant activity loss on labeling and isolation. The slight activity in PNA+ cells may have been contaminants. The PNA- Ly 2+ subset formed larger clones than the PNA- Ly 2- subset. CTL-p frequency was 1 in 5 for PNA- Ly 2+ and 1 in 400 for PNA- Ly 2-. The few cytotoxic clones derived from the Ly 2- cells appeared to be genuine and not a result of contamination with Ly 2+ cells. Thus although both Ly subsets of medullary-type thymocytes were able to proliferate, the Ly 2+ subset contributed almost all of the cytotoxic activity of the unfractionated thymocytes. Medullary-type thymocytes display an Ly phenotype development and a level of functional maturation approaching that of peripheral T cells.
对于具有不同Ly表型的小鼠胸腺和外周T细胞亚群，测定了所有能够增殖的T细胞前体（PTL-p）以及细胞毒性T细胞克隆前体（CTL-p）的频率。使用了高克隆效率、伴刀豆球蛋白A（Con A）和生长因子驱动的极限稀释培养系统。采用凝集素介导的非特异性裂解读数来检测细胞毒性克隆。这种方法提供了一份不考虑特异性的功能细胞总体分布情况的资产负债表。在用单克隆抗Thy 1和抗Ly 2抗体进行双色染色后，通过荧光激活细胞分选（FACS）分离脾T淋巴细胞亚群。Ly 1+2-和Ly 1+2%亚群均通过克隆增殖做出反应，但细胞毒性活性几乎完全局限于源自Ly 1+2%的克隆。在用花生凝集素（PNA）和单克隆抗Ly 2抗体进行双色染色后，通过FACS分离胸腺细胞的四个亚类。它们分别是PNA+Ly 2+、PNA+ Ly 2-、PNA- Ly 2+和PNA- Ly 2-，分别占胸腺细胞总数的80%、5%、5%和10%。它们各自的PTL-p频率分别为1/333、1/200、1/5.3和1/3.2，这些值包括标记和分离过程中的显著活性损失。PNA+细胞中的轻微活性可能是污染物造成的。PNA- Ly 2+亚群形成的克隆比PNA- Ly 2-亚群的大。PNA- Ly 2+的CTL-p频率为1/5，PNA- Ly 2-的为1/400。源自Ly 2-细胞的少数细胞毒性克隆似乎是真实的，并非Ly 2+细胞污染的结果。因此，尽管髓质型胸腺细胞的两个Ly亚群都能够增殖，但Ly 2+亚群几乎贡献了未分级胸腺细胞的所有细胞毒性活性。髓质型胸腺细胞表现出Ly表型发育以及接近外周T细胞的功能成熟水平。
