Enucleated cytotoxic T lymphocytes bind specifically to target cells in vitro

We report the isolation of enucleated particles (cytoplasts) from mouse cytotoxic T lymphocytes (CTLs) which, like intact CTLs, can specifically bind to target cells (TCs) in vitro. CTL-enriched populations were enucleated by centrifugation on a Ficoll density gradient at 31 C in the presence of cytochalasin B. The resulting cytoplasts which consist of 99% of enucleated particles retained about 40% of the nucleated cells' protein but less than 1% of their DNA. The presence of cell surface membrane antigens Thy 1.2 and H-2 on the cytoplasts indicated that their surface membranes originated from the plasma membranes of the intact cells. Electron microscopy of cytoplasts revealed two types of particles, one (type a) contained cytoplasmic organelles such as mitochondria, Golgi apparatus, microfilaments, and microvilli, while the second (type b) contained little cytoplasm and no microfilaments or microvilli. The evidence presented suggests that only type a cytoplasts can bind specifically to TCs. The specific binding to TCs by type a and not type b cytoplasts shows that while the CTL nucleus or nucleus-associated structures affected by enucleation are not essential for specific CTL receptor activity, cytoskeletal structures and activities related to these structures must be preserved.