We developed phospho-ERK1/2 ELISA for human and rainbow trout liver cells, employing HepG2 and RTL-W1 cell lines as models. The assay was applied to detect changes in ERK1/2 activity for nine chemicals, added over a wide concentration range and time points. Cell viability was measured to separate ERK1/2 regulation from cytotoxicity. Perfluorooctane sulfonate and carbendazim did not change ERK1/2 activity; influence on ERK1/2 due to cytotoxicity was indicated for tributyltin and cypermethrin. Mancozeb, benzo[a]pyrene, and bisphenol A stimulated ERK1/2 up to ∼2- (HepG2) and 1.5 (RTL-W1)-fold, though the kinetics differed between chemicals and cell lines. Bisphenol A and benzo[a]pyrene were the most potent concentration-wise, altering ERK1/2 activity in pM (HepG2) to nM (RTL-W1) range. While atrazine and ibuprofen increased ERK1/2 activity by ∼2-fold in HepG2, they did not initiate an appreciable response in RTL-W1. This assay proved to be a sensitive, medium- to high-throughput tool for detecting unrecognized ERK1/2-disrupting chemicals.

我们使用 HepG2 和 RTL-W1 细胞系作为模型开发了用于人和虹鳟鱼肝细胞的磷酸化 ERK1/2 ELISA。该测定用于检测九种化学物质的 ERK1/2 活性变化，添加到很宽的浓度范围和时间点。测量细胞活力以将 ERK1/2 调节与细胞毒性分开。全氟辛烷磺酸盐和多菌灵不改变 ERK1/2 活性；表明三丁基锡和氯氰菊酯由于细胞毒性而对 ERK1/2 产生影响。代森锰锌、苯并[ a ]芘和双酚 A 将 ERK1/2 刺激至 2-（HepG2）和 1.5（RTL-W1）-倍，尽管化学物质和细胞系之间的动力学不同。双酚A和苯并[ a]芘是最有效的浓度，在 pM (HepG2) 到 nM (RTL-W1) 范围内改变 ERK1/2 活性。虽然莠去津和布洛芬在 HepG2 中将 ERK1/2 活性提高了约 2 倍，但它们在 RTL-W1 中并未引发明显反应。该测定被证明是一种灵敏的中高通量工具，用于检测无法识别的 ERK1/2 干扰化学物质。