A practical label-free method for the rapid determination of small-molecule critical micelle concentration (CMC) using a fixed-angle light-scattering technique is described. Change in 90° light scattering at a fixed wavelength of incident radiation with increasing bacterial quorum molecule concentration and the observation of a break point is used to determine CMC. In our study, this technique is utilized to investigate the aqueous CMC of previously uncharacterized Pseudomonas aeruginosa quorum sensing signaling molecules (QSSM) belonging to the n-acylhomoserine lactone and 2-alkyl-4-quinolone classes. Several were found to form micelles within a physiologically relevant concentration range and potential roles of these micelles as QSSM transporters are discussed. The influence of temperature and the presence of biological membranes or serum proteins on QSSM CMC are also investigated and evidence is obtained to suggest the QSSMs studied are capable of both membrane and serum protein interaction. This demonstrates that the fixed-angle light-scattering technique outlined can be used simply and rapidly to determine small-molecule CMC under a variety of conditions.