The mechanism of antiviral activity of the CD4-derived peptide 75-99 was compared with that of sulfated polysaccharides. A set of peptides representing all the high positive charge density regions of gp120 and gp41 was used to determine whether electrostatic interactions occur between these negatively charged agents and positively charged HIV envelope fragments. Synthetic peptide AZ2, amino acids 75-99 from V1 CD4, KIEDSDTYIC(Acm)-EVEDQKEEVQLLVFG, and dextran sulfate 500,000 (DS 500) were used as inhibitory agents of antibody binding in ELISA using: (1) anti-peptide rabbit antibodies; (2) sera from HIV infected persons. Peptide AZ2 and DS were both shown to block antibody binding to peptide (301-323) from the principal neutralizing domain (PND) and peptide (495-516) from the gp120 C-terminus. The blocking concns were 1-2 micrograms/ml for DS and 125-250 micrograms/ml for AZ2. The ELISA system based on rabbit anti-peptide antibodies was less sensitive than that based on positive human sera. Chemical modification of lysine epsilon-amino groups of these peptides resulted in complete failure to bind either DS or AZ2. A correlation was found between the inhibitory activities of a number of sulfated polysaccharides in a syncytium formation assay and in peptide ELISA. The mechanism of direct interactions of specific regions of gp120 with the CDR3-like region of CD4 is proposed.