A SHORT-DURATION POLYETHYLENE-GLYCOL FUSION TECHNIQUE FOR INCREASING PRODUCTION OF MONOCLONAL ANTIBODY-SECRETING HYBRIDOMAS
A SHORT-DURATION POLYETHYLENE-GLYCOL FUSION TECHNIQUE FOR INCREASING PRODUCTION OF MONOCLONAL ANTIBODY-SECRETING HYBRIDOMAS
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DOI：
10.1016/0022-1759(85)90207-8
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发表时间：
1985-01-01
影响因子：
2.2
通讯作者：
LANE, RD
中科院分区：
文献类型：
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作者：
LANE, RD
评价淋巴细胞-骨髓瘤细胞混合物作用于FusoGen的时间对总杂交瘤集落产量和分泌单抗的影响。将SP2/0和Fox-NY骨髓瘤细胞与绵羊红细胞免疫的小鼠脾淋巴细胞进行不同时间的融合。最佳融合时间为加入FusoGen(5.0mlKodak1450peg[聚乙二醇]，0.5ml二甲基亚砜，4.5ml磷酸盐缓冲盐水，pH 76.0)到细胞混合物中，时间为45 S，温度为37度。C.通过将混合物逐渐稀释到50毫升的RPMI-1640中来停止融合过程。10min后，将细胞离心，悬浮在含饲养层巨噬细胞的选择性培养液中进行培养。与普通的更长时间的融合技术相比，该程序产生的杂交细胞数量在使用Sp2/0细胞时增加了约5倍，在使用Fox-NY细胞作为融合伙伴时产生的杂交细胞数量增加了30倍。在这两种情况下，几乎所有的孔都含有分泌单抗的杂交瘤克隆。这种高效的融合技术可以最有利地用于生产针对弱免疫原的单抗或减少用更强的免疫原免疫所需的时间。
The exposure period of the lymphocyte-myeloma cell mixture to the fusogen was evaluated for its influence upon the yield of total hybridoma colonies and those which secreted monoclonal antibodies. Sp2/0 and FOX-NY myeloma cells were fused for varying periods with murine splenic lymphocytes immunized with sheep red blood cells. The optimal fusion period consisted of adding the fusogen (5.0 ml Kodak 1450 PeG [polyethylene glycol], 0.5 ml dimethylsulfoxide, and 4.5 ml of phosphate-buffered saline, pH 76.0) to the cell mixture over a 45 s period at 37.degree. C. The fusion process was stopped by gradually diluting the mixture in 50 ml of RPMI-1640. After 10 min, the cells were centrifuged, resuspended in selective medium with feeder macrophages and cultured. In comparison to common, longer fusion techniques, this procedure produces approximately a 5-fold increase in the number of hybrid sproduced when using the Sp2/0 cells and a 30-fold increase in the number of hybrids produced when using the FOX-NY cells as the fusion partner. In both cases, virtually all the wells contain monoclonal antibody-secreting hybridoma colonies. This high efficiency fusion technique can be used most advantageously to produce monoclonal antibodies against weak immunogens or to reduce the time needed for immunization with stronger immunogens.