Production of fluorescein isothiocyanate conjugated mouse monoclonal anti-heparan sulfate proteoglycans for immunologic assay

Abstract

Heparan sulfate proteoglycans (HSPGs) involves significantly in biological activities and acts as the co-receptor of many certain molecules with heparin binding region or some pathogen. Previous study of Assistant Professor Dr. Preeyanat Vongchan, Human liver HSPGs was first isolated from human liver and monoclonal antibodies were produced. 1E4-1C2, one of many clones obtained was proved to specifically reacts with cell membrane molecules of acute myeloid leukemia (AML) by indirect immunofluorescent technique. Since the method is time consumed and requires FITC-conjugated anti-mouse Igs which is expensive, we therefore aimed to prepare fluorescein isothiocyanate (FITC) conjugated MAbs clone 1E4-1C2 to value add of anti-human liver HSPGs. FITC was covalently coupled to primary amines (lysine) of the immunoglobulin. The result showed that F/P ratio of this product is 4.73. Moreover, antibody concentration showing significantly positive result (5%) with HepG2 cell line by direct immunofluorescent is 40 μg/ml. Finally, we could prepared FITC conjugated anti-human liver HSPGs and use to investigate AML by direct immunologic application. Bull Chiang Mai Assoc Med Sci 2009; 42: 50-56.