Carvajal R E, Porras F M, Garcia-Carreño F
Immunol Lett. 1984;8(2):101-6. doi: 10.1016/0165-2478(84)90059-2.
In order to separate, isolate, and determine the number and distribution of the subpopulations of lymphocytes of diverse affinities that are present in an immune response toward a single hapten, anti-trinitrophenyl (TNP) lymphocytes from immunized animals were purified by cell chromatography. Non-adherent spleen cells were passed through a column consisting of TNP-substituted polyacrylamide beads. The retained cells were eluted by applying a linear concentration gradient of TNP-lysine. Elution profiles having a limited number of peaks were obtained in all cases. The avidity of the cells in each fraction was measured by inhibition of formation of immune rosettes by free hapten. Results showed that each peak was located along the gradient according to its affinity since there was a direct correlation between the affinity and the concentration of hapten needed for the elution. The cells in each peak appeared to belong to a homogeneous subpopulation as shown by the slope of the curves obtained in the determination of avidity, suggesting that each peak corresponded to one expanded clone.
为了分离、纯化并确定在针对单一半抗原的免疫反应中存在的不同亲和力淋巴细胞亚群的数量和分布，通过细胞色谱法对来自免疫动物的抗三硝基苯（TNP）淋巴细胞进行了纯化。非黏附性脾细胞通过由TNP取代的聚丙烯酰胺珠组成的柱。保留的细胞通过应用TNP-赖氨酸的线性浓度梯度进行洗脱。在所有情况下均获得了具有有限数量峰的洗脱图谱。通过游离半抗原对免疫玫瑰花结形成的抑制作用来测量每个级分中细胞的亲和力。结果表明，由于亲和力与洗脱所需的半抗原浓度之间存在直接相关性，每个峰根据其亲和力沿着梯度定位。如在亲和力测定中获得的曲线斜率所示，每个峰中的细胞似乎属于一个同质亚群，这表明每个峰对应于一个扩增的克隆。
