A novel chitin-binding lectin was isolated from the tubers of a plant Aponogeton natans from the monocot family Aponogetonaceae, designated as ANTL (Aponogeton natans tuber lectin). The lectin agglutinated both untreated and trypsin-treated rabbit erythrocytes, as well as human blood cells of groups A, B and O with different specificities. Lectin activity is inhibited by the oligomers of N-acetylglucosamine. ANTL is a dimeric glycoprotein with molecular weight of ∼66 KDa and has two identical sub-units of 33 KDa. The carbohydrate percent is 8.2% of the total lectin. The lectin was thermo stable up to 50°C with broad pH optima (pH 4–10). ANTL is found to be potent mitogen for normal murine and human lymphocytes at the concentration as low as 1 µg/ml. Cytotoxic studies of the lectin on human U 266 cell lines has revealed that there is 50% decrease in the proliferation. The confirmation of both the hemagglutination and mitogenic proliferation activity suggests that ANTL is a Chitin-binding lectin with diverse functions. The pharmacological relevance of ANTL as a potent mitogen with some cytotoxic effect in certain cell lines are reported for the first time.HighlightsA novel chitin-binding lectin was purified from the tuber extracts of Aponogeton natans (ANTL) in a single step on chitin column by affinity chromatography.ANTL is a dimeric glycoprotein with a molecular weight of ∼66 KDa with high hemagglutination activity towards rabbit erythrocytes.The cytotoxic effect of ATNL on human cell lines U266 has shown 50 % inhibition of their proliferation.ANTL displayed potent mitogenic response towards murine and human peripheral blood mononuclear cells.