Using a fluorescein-conjugated antigenic peptide, peptide-receptive H-2Kb MHC class I molecules were found throughout the secretory pathways of RMA cells and peptide transporter (TAP)-deficient derivative cells (RMA/S). RMA/S cells displayed higher levels of intracellular peptide-receptive molecules, while, surprisingly, RMA cells expressed 3- to 5-fold more cell surface peptide-receptive molecules. Metabolic radiolabeling of Kb-associated oligosaccharides with [1-3H]galactose demonstrated that despite a large difference in the fraction of Kb molecules in native conformation in detergent extracts, Kb transport rates from the trans-Golgi complex to the surfaces of RMA and RMA/S cells were similar. Thus, although considerable numbers of class I alpha chains reach the RMA/S cell surface, they are a less productive source of peptide-receptive molecules than class I molecules synthesized by TAP-expressing RMA cells, suggesting paradoxically that TAP functions to increase the amount of peptide-receptive molecules at the cell surface.