The discovery of Toll-like receptors (TLRs) represented a significant breakthrough that paved the way for the study of host-pathogen interactions in innate immunity. However, there are still major gaps in understanding TLR function, especially the early dynamics of downstream TLR pathways remains less clear. We characterize a label-free optical biosensor-based assay as a powerful method to detect TLR activation in a native and label-free environment and to delineate the dynamics of TLR pathway activation. To gain a deeper insight into the biological processes underlying the ligand-specific signal traces of different LPS chemotypes in various cell types, RNA-seq analysis of transcriptional changes in THP-1 macropahges was performed. After 3 hours of stimulation with LPS E. coli or LPS S. minnesota, THP-1 macrophages showed significant changes in RNA expression compared to the unstimulated control. Overall design: THP-1 macrophages were stimulated with LPS E. coli or LPS S. minnesota for 3 hours. Unstimulated cells served as control. Gene expression profiling analysis was performed using data obtained from RNA-seq of two independent experiments.