Con A lymphoblasts were treated with 2 mM TNBS and then cultured for 24 hr. At the time of treatment, approximately half of the cell membrane H-2 molecules were derivatized with TNP, whereas after culture less than 10% of the H-2 molecules were TNP derivatized. When the TNBS-treated and cultured cells were tested in a cold target competition assay for antigenic activity by using anti-TNP cytotoxic effector cells, it was found that these cells were greatly reduced in their ability to inhibit activity. This lack of activity could be overcome by retreatment of the inhibitor cells with 2 mM TNBS such that these cells now displayed TNP-H-2 molecules and blocking activity in the TNP-CML assay. Although the cultured cells shed 50% of their TNP molecules, it is unlikely that the failure of these cells to block in the TNP-CML assay was due to loss, since cells treated with 0.5 mM TNBS but used without culturing were able to efficiently block in the competition assay. The 0.5 mM TNBS-treated cells displayed an equivalent amount of TNP molecules bound per cell as the cells treated with 2 mM TNBS and cultured for 24 hr.
Thus, the data indicate that after 24 hr of culture of TNBS treated cells, H-2 molecules turnover rapidly so that the TNP-H-2 molecules are replaced. These cells fail to express antigenic activity that can be recognized by cytotoxic anti-TNP effector cells.