Tice D G, Davey F R
Clin Exp Immunol. 1986 Feb;63(2):321-6.
T lymphocyte colonies, arising from phytohemagglutinin (PHA) stimulated mononuclear cells cultured in a semi-solid agar matrix, could be the progeny of single cells (monoclonal) or of multiple cells (polyclonal). We have conducted several studies to determine if these colonies are monoclonal or polyclonal in origin. Normal human peripheral blood mononuclear cells from male-female, HLA-A and B disparate donor pairs were incubated for 18 h in RPMI 1640 containing PHA and fetal calf serum (FCS) and then cultured in a two-layer semi-solid agar system. After 5 days of incubation, the clonality of the colonies was assessed by in situ Y chromatin analysis, and by analysis of HLA-A and B locus antigens. Overlayers were stained with quinicrine dihydrochloride and the number of cells in the T cell colonies with Y chromatin enumerated using fluorescence microscopy. In other studies, colonies were picked from the agar with a capillary pipette and expanded in culture media. After 17 days of culture, cells were harvested and HLA-A and B phenotypes were determined. The results indicate that 87% of the T cell colonies had cells of either male or female origin. In addition, 90% of the colonies possessed HLA-phenotypes of only one donor. We conclude that Y chromatin and HLA analysis of individual colonies from cocultures suggest the monoclonality of T lymphocyte colonies.
由植物血凝素（PHA）刺激、在半固体琼脂基质中培养的单核细胞产生的T淋巴细胞集落，可能是单细胞（单克隆）或多细胞（多克隆）的后代。我们进行了多项研究，以确定这些集落的起源是单克隆还是多克隆。来自男女、HLA - A和B不同供体对的正常人外周血单核细胞，在含有PHA和胎牛血清（FCS）的RPMI 1640中孵育18小时，然后在双层半固体琼脂系统中培养。孵育5天后，通过原位Y染色质分析以及HLA - A和B位点抗原分析来评估集落的克隆性。上层用二盐酸奎宁染色，使用荧光显微镜计数具有Y染色质结构的T细胞集落中的细胞数量。在其他研究中，用毛细吸管从琼脂中挑取集落，并在培养基中扩增。培养17天后，收获细胞并确定HLA - A和B表型。结果表明，87%的T细胞集落中的细胞只有男性或女性来源。此外，90%的集落仅具有一个供体的HLA表型。我们得出结论，对共培养中单个集落的Y染色质和HLA分析表明T淋巴细胞集落具有单克隆性。
