Key parameter optimization using multivariable linear model for the evaluation of the in vitro estrogenic activity assay in T47D cell lines (CXCL-test)

0558418 - MBÚ 2023 RIV US eng J - Journal Article
Linhartová, Lucie - Costet, N. - Pakdel, F. - Cajthaml, Tomáš - Habauzit, D.
Key parameter optimization using multivariable linear model for the evaluation of the in vitro estrogenic activity assay in T47D cell lines (CXCL-test).
Journal of Applied Toxicology. Roč. 42, č. 7 (2022), s. 1121-1136. ISSN 0260-437X. E-ISSN 1099-1263
R&D Projects: GA ČR(CZ) GJ20-14318Y
Institutional support: RVO:61388971
Keywords : genetic instability * luciferase reporter * breast * endocrine * culture * drinking * serum * cytokine CXCL12 * sdf1 * estrogenic activity cellular assay * material variability * multivariate linear model (MLM) * t47d
OECD category: Toxicology
Impact factor: 3.3, year: 2022
Method of publishing: Limited access
https://analyticalsciencejournals.onlinelibrary.wiley.com/doi/10.1002/jat.4280

In comparison with analytical tools, bioassays provide higher sensitivity and more complex evaluation of environmental samples and are indispensable tools for monitoring increasing in anthropogenic pollution. Nevertheless, the disadvantage in cellular assays stems from the material variability used within the assays, and an interlaboratory adaptation does not usually lead to satisfactory test sensitivities. The aim of this study was to evaluate the influence of material variability on CXCL12 secretion by T47D cells, the outcome of the CXCL-test (estrogenic activity assay). For this purpose, the cell line sources, sera suppliers, experimental and seeding media, and the amount of cell/well were tested. The multivariable linear model (MLM), employed as an innovative approach in this field for parameter evaluation, identified that all the tested parameters had significant effects. Knowledge of the contributions of each parameter has permitted step-by-step optimization. The most beneficial approach was seeding 20,000 cells/well directly in treatment medium and using DMEM for the treatment. Great differences in both basal and maximal cytokine secretions among the three tested cell lines and different impacts of each serum were also observed. Altogether, both these biologically based and highly variable inputs were additionally assessed by MLM and a subsequent two-step evaluation, which revealed a lower variability and satisfactory reproducibility of the test. This analysis showed that not only parameter and procedure optimization but also the evaluation methodology must be considered from the perspective of interlaboratory method adaptation. This overall methodology could be applied to all bioanalytical methods for fast multiparameter and accurate analysis.
Permanent Link: http://hdl.handle.net/11104/0332226