Mao S J, France D S
J Immunol Methods. 1984 Dec 31;75(2):309-16. doi: 10.1016/0022-1759(84)90114-5.
The limiting dilution technique is a critical step in the cloning of hybridomas for the preparation of monoclonal antibodies. We have found that culture medium supplemented with human plasma low density lipoproteins (LDL) markedly enhanced the yield of hybridoma clones derived from P3 X 63 Ag or FO mouse myeloma cell lines upon limiting dilution. Such enhancement was dependent on the concentration of LDL employed, being optimal at 1-10 micrograms/ml. At LDL concentrations greater than 20 micrograms/ml, the increase in yield of hybrid clones was not significant. The mechanism by which LDL enhances the yield of hybrid clones was partially elucidated by the demonstration that LDL could increase the DNA synthesis of hybridomas as assessed by [3H]thymidine incorporation. The data suggest that LDL play a role in the proliferation of hybridomas. It also indicates that LDL can be utilized for limiting dilution to increase the yield of desired clones. Since LDL is one of the most abundant lipoprotein fractions (approximately 500 micrograms/ml) in human plasma and the isolation procedure is simple, hybridoma culture medium supplemented with human LDL will prove to be a valuable reagent for investigators currently employing monoclonal antibody technology.
有限稀释技术是用于制备单克隆抗体的杂交瘤克隆过程中的关键步骤。我们发现，在有限稀释时，添加人血浆低密度脂蛋白（LDL）的培养基能显著提高源自P3 X 63 Ag或FO小鼠骨髓瘤细胞系的杂交瘤克隆产量。这种提高取决于所使用的LDL浓度，在1 - 10微克/毫升时最为理想。当LDL浓度大于20微克/毫升时，杂交克隆产量的增加并不显著。通过[3H]胸苷掺入法评估发现LDL可增加杂交瘤的DNA合成，这部分阐明了LDL提高杂交克隆产量的机制。数据表明LDL在杂交瘤增殖中发挥作用。这也表明LDL可用于有限稀释以提高所需克隆的产量。由于LDL是人类血浆中最丰富的脂蛋白组分之一（约500微克/毫升）且分离程序简单，添加人LDL的杂交瘤培养基将被证明是目前采用单克隆抗体技术的研究人员的一种有价值的试剂。
