Addition of 2 to 10 micrograms of wheat germ agglutinin (WGA), a lectin from Triticum vulgaris specific for N-acetyl-D-glucosamine, per ml to suspensions of mouse fibroblasts (L cells) blocked the attachment of 14C-labeled Chlamydia psittaci 6BC to the L-cell surface. WGA and strain 6BC competed for similar sites on L cells, but once bound, one was not replaced by the other. N-Acetyl-D-glucosamine, but not other monosaccharides of related structure, antagonized the blocking action of WGA. Lectins with specificities other than that of WGA prevented chlamydial attachment only at much higher concentrations or not at all. Exposure of L cells to trypsin and to high multiplicities of strain 6BC decreased the amount of subsequently added 3H-labeled WGA that was bound by these cells. WGA also blocked the attachment of strain 6BC to other established cell lines of murine, simian, and human origin. A lymphogranuloma venereum strain (440L) of C. trachomatis was just as sensitive to the blocking action of WGA as was strain 6BC. It appears that the attachment of both C. psittaci and C. trachomatis to host cells of diverse origin involves an N-acetyl-D-glucosamine-containing entity that binds WGA with high affinity.