Abstract We have previously shown that antibody responses are inhibited following administration of κ-opioid agonists. We found that the inhibition was blocked by either naloxone or the κ-selective antagonist norbinaltorphimine. This inhibitory activity is apparent after short-term treatment with the κ-opioid agonist. In an attempt to identify the cell populations which serve as the target for this immunosuppressive effect, we have carried out cell fractionation analyses to generate isolated T cells and macrophages. Using multiple cell fractionation methods, we have determined that short-term treatment of either T cells or macrophages with the κ-opioid agonist U50,488H results in significant inhibition of in vitro antibody responses. We also find that the inhibition of both T cell and macrophage activity can be blocked by naloxone. These studies demonstrate that resting T cells and macrophages express κ-opioid receptors and exhibit significant opioid responsiveness prior to activation by antigen.