Human Lymphoblastoid Cell Lines. III. Co-cultivation Technique for Establishment of New Lines2

Abstract

Twenty-five new human lymphoblastoid cell lines were grown by a cocultivation method in which lethally irradiated cells from an established line were mixed in vitro with fresh peripheral blood leukocytes. Only material from Epstein-Barr virus (EBV)-positive lines had the property of inducing long-term growth in fresh leukocytes. The property persisted for up to 7 days (in one case 9 days) after irradiation. The likelihood of initiating a new cell line was not affected by the number of irradiated cells/5 ml mixed culture (within the range 0.5 × 10⁶ to 5 × 10⁶) but was significantly improved when the number of fresh leukocytes exceeded 3 × 10⁶. Placental blood leukocytes gave rise to new cell lines more readily than those from children or adults. Electron microscopy revealed EBV in only 8 of 23 lines examined. It is suggested that occult infection was present in the others. All but 1 new line had a diploid karyotype, and no evidence was found for a specific effect of EBV infection on the chromosomes of human lymphoblastoid cells.