Endocrine Abstracts

The chemically induced disturbance during the neurodevelopment stage could cause a serious disease. In vivo study is useful to discriminate against neurotoxic substances but is time-consuming and ethically problematic. So, we have previously established, developmental neurotoxicity test (DNT) in vitro method using Sox1-GFP. In this study we aimed to increase the predictability of discriminant function. The equation was statistically improved using thirty additional chemicals. Concomitantly, we conduct reproducibility assessment; inter- and intra- studies were carried out for fifteen chemicals by three experimenter and three independent laboratories. In this study, we correlated our prediction model with wider range of chemicals; 20 toxicants and 10 non-toxicants data was newly added to secure analytical accuracy. To establish a discriminant function, two factors, half inhibition concentration for cell viability (IC50) and half inhibition concentration for neuro-sphere area (ID50), were used. IC50 value was obtained by performing CCK-8 assay after chemical treatment on undifferentiated cells for 2 days. ID50 value was obtained by measuring both GFP intensity and neuro-sphere areas of differentiated cells as neural form which cell was exposed to chemical for 4 days. From the inter result obtained using the stated discriminant function gave 100% accuracy and from intra test it gave 96% accuracy. Inter- and intra- experiments were able to classify chemicals as toxic or non-toxic very equally. Conclusively we have established animal alternative DNT and showed that this test is a transferable, reproducible and accurate method. Therefore, we suggest that this method will be able to be a standardizing method as animal alternative. Keyword: Developmental neurotoxicity test, in vitro, Sox1-GFP cell