Chemical modification studies with group specific reagents indicated that the imidazole side chains of histidine residues are involved in the carbohydrate-binding activity of Trichosanthes cucumerina seed lectin (TCSL). A total of 9.8 (±1.8) histidine residues could be modified by reaction with diethyl pyrocarbonate when the reaction was carried out for 2 hours, which resulted in a total loss in the carbohydrate binding and hemagglutinating activities of the lectin. Reversing the modification by treating the histidine-modified protein with hydroxylamine resulted in a complete recovery of the activity. Presence of the specific sugar (0.2 M galactose) afforded a partial protection in that only 4.6 His residues could be modified in the same period of time. In the presence of 6 M guanidinium hydrochloride, 15.8 (±1.5) His residues were modified. The histidine-modified lectin cross-reacted with rabbit anti-TCSL antiserum, indicating that the conformation of the modified lectin is unaltered and that the loss of activity is not a consequence of structural changes. Modification of the side chains of lysine, tyrosine and cysteine residues did not affect the haemagglutination activity of the lectin. The tryptophan residues of native TCSL could not be modified by N-bromosuccinimide. However, modification with this reagent in the presence of 8 M urea indicated that there are 4.6 (±0.4) Trp residues in the dimeric lectin.