The immunoregulatory normal plasma lipoprotein subclass, termed LDL-In, suppresses a number of immune responses in vitro including peripheral blood lymphocyte phytohemagglutinin-stimulated 3H-thymidine uptake, pokeweed mitogen-driven immunoglobulin synthesis, and the generation of cytolytic T cells. In vivo parallels have also been demonstrated. Plasma lipoproteins represent noncovalent complexes of lipid and apolipoprotein chains. Since free fatty acids have been demonstrated to suppress certain lymphocyte responses including mitogen-stimulated 3H-thymidine uptake, we have examined the hypothesis that the free fatty acids associated with LDL-In particles are the biologically active constituents. However, the dose-dependence and the kinetics of LDL-In-mediated suppression were distinctly different from those observed for a number of free fatty acids. In addition, when greater than 92% of the endogenous free fatty acids of LDL-In were removed by incubation in the presence of delipidated albumin, no differences were observed between this free fatty acid depleted LDL-In and native LDL-In with respect to suppressive activity for lymphocyte stimulation or binding to the lymphocyte LDL-In receptor. These observations indicate that the interaction of the LDL-In particle with the lymphocyte receptor and the suppression of lymphocyte function are independent of the free fatty acid content of this bioregulatory lipoprotein, thus focusing attention on the apolipoprotein chains or the phospholipids and glycolipids of this minor plasma lipoprotein.