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前列腺素E2(prostaglandins E2,PGE2)是参与炎症、疼痛等多种生理病理机制过程的活性物质。该文在脂多糖(lipopolysaccharide,LPS)诱导小鼠巨噬细胞RAW264.7产生PGE2的模型上,利用均相时间分辨荧光(homogeneous time-resolved fluorescence,HTRF)方法结合全自动化筛选操作系统,建立检测PGE2含量的高通量模型,选用公司中药物质基础库内的5 121个中药成分为检测样品,用于筛选PGE2抑制剂。该模型选择细胞铺板密度为每孔2 000个细胞;阳性化合物的IC50平均值为(7.3±0.1)μmol;检测板的Z’因子均大于0.5,平均值为0.7;5 121个样品中有228个样品在检测浓度对PEG2产生的抑制率大于50%,23个样品对PEG2产生的抑制作用大于80%。该模型数据稳定性和准确性均达到预期标准,表明筛选结果可靠真实;自动化筛选系统的引入,令该模型具有快速,微量,高效等优势,日平均筛选量达到14 000数据点以上,为抗炎镇痛相关新药发现和中药有效物质成分的前期快速筛选提供了新的模型。
Prostaglandins E2 (PGE2) is an active substance that participates in many physiological and pathological processes such as inflammation and pain. In this study, lipopolysaccharide (LPS) -induced macrophage RAW264.7-induced PGE2 was detected by a homogeneous time-resolved fluorescence (HTRF) combined with a fully automated screening system PGE2 content of the high-throughput model, the selection of the company’s traditional Chinese medicine base of 5 121 Chinese herbal ingredients as test samples for screening PGE2 inhibitors. In this model, cell plating density was selected to be 2 000 cells per well; the IC50 values for the positive compounds were (7.3 ± 0.1) μmol; the Z ’factors for the assay plate were greater than 0.5 with an average of 0.7; 228 of the 5 121 samples The inhibition rate of PEG2 by these samples was more than 50% at the concentration tested, and the inhibitory effect of 23 samples on PEG2 was more than 80%. The stability and accuracy of the model data meet the expected standards, indicating that the screening results are reliable and true. The introduction of automated screening system makes the model fast, trace and efficient with the average daily screening of more than 14 000 data points A new model has been provided for the rapid pre-screening of the discovery of new drugs related to analgesia and the effective components of traditional Chinese medicine.