牛视网膜提取物对人-鼠杂合细胞抗体产生的影响

Effects of Bovine Retinal Extract on Production of Human IgM in the Culture of Human-Mouse Hybridoma Cells

目的：探讨牛视网膜提取物对人－鼠杂合细胞抗体产生的影响。方法：在ＰＲＭＩ１６４０完全培养基（ＣＭ）中加入２０ｍｇ／Ｌ牛视网膜提取蛋白，配成牛视网膜培养基（ＢＲＥＭ）。用ＭＴＴ测定、细胞克隆技术、夹心ＥＬＩＳＡ方法和染色体分析，比较人－鼠杂合细胞Ｇ１２在两种培养条件下的细胞活力、克隆效率、人ＩｇＭ分泌和人染色体阳性细胞比率之间的差异。结果：用ＢＲＥＭ培养的Ｇ１２细胞活力高于用ＣＭ培养（Ｐ＜００５）。用ＢＲＥＭ培养的Ｇ１２细胞克隆中分泌人ＩｇＭ者占１２／２４，用ＣＭ培养的克隆只占２／４７。Ｇ１２细胞传代培养３个月后，用ＢＲＥＭ培养的上清人ＩｇＭ水平Ａ４９０为０３３５±００５０，用ＣＭ培养的上清为００７０±００２７（Ｐ＜００５）。前者含人染色体阳性的细胞为２０％，后者２５％，两者间没有差异（Ｐ＞００５）。结论：添加牛视网膜提取物的培养基可以提高人－鼠杂合细胞的活力和人ＩｇＭ抗体的分泌能力。维持人源性ＩｇＭ持续分泌的条件不仅取决于杂合细胞中人染色体的存在和稳定，还有其它未知因素在起作用，牛视网提取蛋白可以提供这方面的需要。

Objective: To study the effects of bovine retinal extract (BRE) on production of human IgM (hIgM) in the culture of human mouse hybridoma cells named G12 . Methods: BRE medium (BREM) was prepared by supplementing 20mg/L protein from BRE to the complete RPMI 1640 medium (CM). The cell viability plating efficiency, the levels of hIgM in supernatant and the rate of human chromosome positive cells in both culture were detecvted with MTT assay, clone technique, double antibody sandwich ELISA and chromosome analysis method respectively. Results: The viability of G12 cell; in BREM was higher than that in CM(P<0 05). In the growth at single cell level, 12/24 of clone secreting hIgM on BREM was obtained, by contrast 2/47 in CM. After generating the cells for 3 months, hIgM levels (A 490  ) of both supernatant (BREM G12 and CM G12) were 0 335±0 050 and 0 070±0 027(P<0 05). 20% of G12 cells in BREM and 25% of G12 cells in CM presented human chromosones (P>0 05).Conclusion: The G12 cells cultured in BRME showed higher cell viability and the ability of secreting hIgM than those in CM. Besides the stability of human chromosome presented in G12 cells, there are some factors in BREM, which could be responsible to the production of hIgM in culture of G12 human mouse hybridoma cells.