U937 cells attach to the RGDS-containing 80-kD fragment of fibronectin (Fn). The present report examined whether these cells recognize other domains of Fn. U937 cells attach to a 38-kD fragment derived from the A chain of Fn, which includes the Hep II domain and most of the alternatively spliced IIICS region. U937 did not bind to a 58-kD fragment derived from the B chain (which lacks IIICS) and has the Hep II site. They also did not bind to a 31-kD COOH-terminal fibrin-binding fragment or to a 29-kD fragment containing the Hep I domain. Cell adhesion to the 38-kD fragment was not inhibited by the 80-kD fragment, by GRGDSPC synthetic peptides, or by a mAb directed to the RGDS-containing domain of Fn. Attachment was completely inhibited by the 38-kD fragment and by the synthetic peptide CS-1, comprising the first 25 amino acid residues of IIICS. These results indicate that U937 cells interact with two sites of Fn, the RGDS-containing region, and the IIICS region.