IC50 values vary in reproducibility and trustworthiness across different experimental assays.
Evidence from multiple studies indicates that IC50 values and assay reproducibility fluctuate depending on the specific experimental assay, readout method, and substrate used.
The retrieved papers consistently demonstrate that IC50 measurements and assay performance vary across different platforms, substrates, and experimental designs, directly supporting the claim.
A. van Tonder, A. Joubert, D. Cromarty. Limitations of the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay when compared to three commonly used cell enumeration assays. 2015. https://doi.org/10.1186/s13104-015-1000-8
The study demonstrates that different cell enumeration assays (such as MTT, NRU, RES, and SRB) exhibit varying degrees of variability and reproducibility when determining IC50 values.
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Shcherbakov DN, Mordvinova ED, Trufanov VO, Volkova NV, Meshkova YV, Marenina MK, Zaykovskaya AV, Volosnikova EA, Borisevich SS, Belenkaya SV. Cell-Based Luciferase Assay for Testing SARS-CoV-2 3CL Protease Inhibitors.. 2026. https://doi.org/10.3390/bios16050253
The research shows that IC50 values obtained for inhibitors differ significantly between cell-based and cell-free assays due to cellular barriers.
Rimmer MA, Wang J, Ranathunge TA, Shi Z, Li Y, Lee D, Chai SC, Lin W, Twarog NR, Ansari AZ, Shelat AA, Young BM, Chen T, Yang L. Label-Free High-Throughput Screening of CYP3A4 Inhibitors Using Acoustic Ejection Mass Spectrometry.. 2026. https://doi.org/10.1021/acs.analchem.5c08059
Findings confirm that both the assay method and the chosen substrate influence which inhibitor hits are detected and their measured potency.
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