【摘要】 用人骨髓瘤细胞株和人脾脏细胞在PEG促进下进行人—人杂交,育选出能分泌抗链球菌溶血素O单克隆抗体杂交瘤S₁—19和S₁—22两株,共传了50多代,历时五个月,单克隆抗体分泌稳定,抗体为IgG型,杂交瘤染色体为68—76条而亲代细胞株仅为43条,证实杂交瘤是成功的。更多还原

【Abstract】 Two human-human hybrid cell lines were established by fusion of human spleen cells and human plasma cells (LICR-LON-HMy2) with PEG (M. W. 1000). For fusion the HMy2 cells were subcultured to the density of optimun , growth daily for two days beforehand. 6 × 107 HMy2 cells and 2.4 × 108 immunized spleen cells were mixed together, then centrifuged for 1000 Rpm/5 minutes. The pellet was resuspended with 10ml serum-free RPMI 1640 and washed three times.All fusions were carried out in 37% PEG (Polyethylene glycol) at room temperature. The cells were seeded at a concentration of 4.5× 105/well in 24-well plates (Costar) in HT medium with 15% calf serum. Hybrid selection in HAT medium was started 24-48 hours after fusion by exchanging half of the HT medium with 2 × HAT medium. Half of the HAT medium was renewed every 3 days in the first 3 weeks after fusion (Therefore the HAT medium contained 10-4 M hypoxanthine, 2 × 10-8 M amino pyterine and 2 ×10-6 M thymidine, all Sigma Chemical Co. USA). The HAT medium was then gradually replaced by HT medium every 4-6 days during the last week. The hybrid cells have been screened for monoclonal antibodies specific to streptolysin O by three times limited dilution with human origin feeder cells (PBC or thymus cells) to confirm the specific antibody positive clones.Some other biological characteristics of these two hybridoma cell lines rramed S1-19 and S1-22 were checked, the karyotype chromosomes ranged 58-76, and the specific antibodies belong to human IgG.Despite the titers of the supernatant of the hybrid cell lines culture were very low, all the data have shown that the human hybridoms have been producing monoclonal antibodies at least for more than 5 months.更多还原