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Bacterial Expression of a Human Monoclonal Antibody-Alkaline Phosphatase Conjugate Specific for Entamoeba histolytica

Departments of Infectious Diseases,¹ Molecular Life Sciences, Tokai University School of Medicine, Isehara, Kanagawa 259-1193,² Department of Tropical Medicine and Parasitology, School of Medicine, Keio University, Shinjuku-ku, Tokyo 160-8582, Japan³

Received 1 August 2003/ Returned for modification 14 September 2003/ Accepted 2 October 2003

We previously produced human monoclonal antibody Fab fragments specific to Entamoeba histolytica in Escherichia coli. In order to use these Fab fragments for diagnostic purposes, an expression vector to produce a fusion protein of Fab and alkaline phosphatase (PhoA) in E. coli was designed and constructed. The E. coli PhoA gene was fused to the 3' terminus of the gene encoding the heavy-chain Fd region. The kappa and Fd genes from a previously prepared antibody clone, CP33, which is specific for the 260-kDa lectin of E. histolytica, were used as human antibody genes. When the fusion protein of CP33 and PhoA was incubated with paraformaldehyde-fixed trophozoites of E. histolytica and developed with a substrate, the trophozoites appeared to be stained. These results demonstrate the feasibility of bacterial expression of a human monoclonal antibody-PhoA conjugate specific for E. histolytica and that the antibody can be used to detect E. histolytica antigen without the use of chemically conjugated secondary antibodies.

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