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用快速自动比色微量分析法检测巨噬细胞条件培养基(MΦCM)对体外培养的生后7d SD大鼠小脑皮质神经元的作用。结果表明,MΦCM内分子量大于30kD的组分(加入量40μl/孔)对神经元(细胞密度1×10~5/孔)具有明显的神经营养活性,与对照组相比有显著性差异(P<0.01)。盖片培养法证明,此组分还具有促神经元突起生长的作用。另外,把MΦCM进行Sephadex G-100凝胶层析及生物活性鉴定,获得具有神经营养活性的第一峰洗脱液。降此洗脱液及上述MΦCM分子量大于30kD的组分经SDS-聚丙烯酰胺梯度凝胶电泳分析,证明MΦCM中含有支持神经元生存和增强其活性作用的成分,是一种分子量为97.9kD的蛋白质。
The effect of macrophage conditioned medium (MΦCM) on cultured rat cerebellar cortical neurons cultured in vitro for 7 days was detected by rapid automated colorimetric assay. The results showed that the components of MΦCM with the molecular weight greater than 30kD (40μl / well) had obvious neurotrophic activity on neurons (cell density 1 × 10-5 / well), which was significantly different from the control group (P <0.01). Cover culture method to prove that this component also has the role of promoting neurite outgrowth. In addition, MΦCM was subjected to Sephadex G-100 gel chromatography and bioassay to obtain the first peak eluate with neurotrophic activity. This eluate and the above MΦCM molecular weight greater than 30kD components by SDS-polyacrylamide gradient gel electrophoresis analysis to prove MΦCM contains neurons to support the survival and enhance its activity of the component is a molecular weight of 97.9kD protein.