Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen associated with worsening disease outcomes in cystic fibrosis (CF) patients. P. aeruginosa uses quorum sensing (QS), a cell-cell signaling system, to control expression of a variety of genes including virulence factors. In P. aeruginosa, QS is mediated in part by acyl-homoserine lactone (AHL) signals that can diffuse in and out of cells. Once AHLs accumulate, they bind to a receptor regulator that activates gene transcription. P. aeruginosa has two complete AHL QS systems, LasI-LasR and RhlI-RhlR. The two systems are arranged in a hierarchy, with the las system controlling the rhl system. QS activation in P. aeruginosa is restrained by cellular proteins that dampen the QS response. These proteins, known as “anti-activators”, attenuate QS by preventing receptor activation. Three anti-activator proteins, QscR, QslA, and QteE, have been identified in P. aeruginosa. These anti-activator proteins have additive, overlapping roles in repressing expression of QS gene products in laboratory strains but their role in the QS dynamics of CF isolates is still unclear. This project used standard molecular cloning techniques to delete or overexpress anti-activator genes in a selection of clinical isolates from CF patients. A reporter plasmid with a fluorescent marker was used to track the activity of LasR and RhlR. These experiments were used to quantify differences in QS-controlled gene activation. To test the hypothesis that anti-activators decrease the amount of LasR in the cell, Western blots were used to assess the cellular levels of QS receptors. In strains with deleted anti-activator genes, LasR levels were higher and induction was earlier. Additional tests for phenotypes controlled by QS, such as protease and pyocyanin production, were also performed. Future research should focus on evaluating these effects in additional CF isolates.