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Fig. 3 Secondary Ca2+ flux assay confirmation. A, The Venn diagram shows the secondary Ca2+ flux assay confirmation results. B, The heat map shows the half maximum effective value (EC50) distribution of all 45 compounds identified and confirmed from secondary assays based on the self-organizing map (SOM) algorithm. The left panel shows the data from the HEK293-GnRHR cell line, and the right panel shows the data from the HEK293-KISS1R cell line. Red indicates a compound with a low EC50 and high potency, and blue indicates a compound with a high EC50 and low potency. Dark blue indicates a compound was inactive in the tested cell line. Compounds can be separated into 3 groups based on their EC50 in each cell line through the SOM algorithm, as indicated by the bold black line. A representative compound for each group and its concentration-response curve are shown in the middle panel.