Dierickx Paul
Instituut voor Volksgezondheid, Afdeling Toxikologie, Laboratorium Biochemische Toxikologie, Brussels, Belgium.
Altern Lab Anim. 2005 Jun;33(3):207-13. doi: 10.1177/026119290503300304.
In our previously described Hep G2/24-hour/total protein assay, protein levels were measured by using the Lowry method. This assay was the best acute in vitro assay for the prediction of human toxicity within the Multicentre Evaluation of In Vitro Cytotoxicity (MEIC) study. In order to increase the MEIC data-base with a wider range of chemicals, we were interested in introducing the more practical 3-(4-carboxybenzoyl)- quinoline-2-carboxaldehyde (CBQCA) method for the quantification of the total protein content. Therefore, we investigated whether the same good results for the prediction of acute human toxicity would be obtained with the CBQCA method. The cells were treated for 24 hours, then cytotoxicity was determined by measuring the total protein content with CBQCA. The results were quantified by using the PI50c: the concentration (in mM) of test compound required to reduce the total protein content measured with the CBQCA-method by 50% as compared to the control cells. The results were compared with the PI50, the corresponding value when the Lowry method was used. A relatively low correlation was observed between PI50 and PI50c, reflecting the large and unexpected, differences when using the two protein assays. However, when comparing the log PI50c with the human toxicity, a correlation coefficient of r(2) = 0.761 (n = 44) was obtained for exactly the same series of MEIC chemicals. This value is clearly higher than that for the Lowry method (r(2) = 0.695). Compared to the Lowry method originally used, the Hep G2/24-hour/CBQCA total protein assay has the additional important advantage that it can be very easily adapted for large-scale analyses with robotic systems, including the on-line calculation of the results.
在我们之前描述的Hep G2/24小时/总蛋白测定中，蛋白质水平采用洛氏法进行测量。该测定是体外细胞毒性多中心评估（MEIC）研究中预测人体毒性的最佳急性体外测定方法。为了用更广泛的化学品增加MEIC数据库，我们有兴趣引入更实用的3-(4-羧基苯甲酰基)-喹啉-2-甲醛（CBQCA）方法来定量总蛋白含量。因此，我们研究了使用CBQCA方法是否能获得与预测急性人体毒性相同的良好结果。将细胞处理培养24小时，然后通过用CBQCA测量总蛋白含量来确定细胞毒性。结果通过PI50c进行量化：与对照细胞相比，使CBQCA法测得的总蛋白含量降低50%所需的受试化合物浓度（以mM为单位）。将结果与PI50进行比较，PI50是使用洛氏法时的相应值。观察到PI50和PI50c之间的相关性相对较低，这反映了使用两种蛋白质测定方法时存在的巨大且意外的差异。然而，当将log PI50c与人体毒性进行比较时，对于完全相同系列的MEIC化学品，获得的相关系数r(2) = 0.761（n = 44）。该值明显高于洛氏法的相关系数（r(2) = 0.695）。与最初使用的洛氏法相比，Hep G2/24小时/CBQCA总蛋白测定具有另一个重要优势，即它可以非常容易地适用于使用机器人系统进行大规模分析，包括结果的在线计算。
