Caspi R R, Rozenszajn L A, Gothelf Y, Pergamenikov-Litvak T, Avtalion R R
Dev Comp Immunol. 1982 Fall;6(4):683-92.
Lymphocytes from peripheral blood of carp proliferate in a clonal culture in soft agar, in the presence of phytohemagglutinin, generating several morphologically distinct types of colonies. Cells from colonies developing on the surface of the agar (surface colonies) and cells from colonies developing within the agar (agar colonies) were studied. Several differences were found between cells from the two types of colonies with respect to morphology, ultrastructure and the distribution of cytoplasmic determinants antigenically related to serum immunoglobulin. Colonies were quantitated as a function of the number of cells seeded, in primary cultures of peripheral blood leukocytes and in secondary (replated) cultures of isolated surface colony cells. The numbers of surface colonies and agar colonies in the two systems were comparable. Preferential formation of surface over agar colonies was noted, and there was an initial concentration of cells (individual for each fish) which resulted in optimal colony growth. This method was found to be suitable for isolating highly homogeneous subpopulations of PHA-responsive lymphocytes, which could subsequently be further expanded in liquid culture. A requirement for an exogenously produced growth factor (possibly similar to mammalian Interleukin 2) in the maintenance of long-term clonal cultures is suggested.
鲤鱼外周血淋巴细胞在软琼脂中、在植物血凝素存在的情况下进行克隆培养时会增殖，产生几种形态上不同类型的集落。对在琼脂表面形成的集落（表面集落）中的细胞和在琼脂内部形成的集落（琼脂集落）中的细胞进行了研究。在形态、超微结构以及与血清免疫球蛋白抗原相关的细胞质决定簇的分布方面，发现这两种集落来源的细胞存在若干差异。在原代外周血白细胞培养物以及分离的表面集落细胞的传代（再接种）培养物中，将集落作为接种细胞数量的函数进行定量。两个系统中表面集落和琼脂集落的数量相当。观察到表面集落比琼脂集落优先形成，并且存在一个初始细胞浓度（每条鱼个体不同），该浓度可导致集落的最佳生长。发现这种方法适用于分离对PHA有反应的淋巴细胞的高度均一亚群，这些亚群随后可在液体培养中进一步扩增。提示在维持长期克隆培养中需要一种外源性产生的生长因子（可能类似于哺乳动物白细胞介素2）。
