Bacterial expression of a human monoclonal antibody-alkaline phosphatase conjugate specific for Entamoeba histolytica

• Published in: Clinical and diagnostic laboratory immunology Vol. 11; no. 1; pp. 216 - 218
• Main Authors: Tachibana, Hiroshi, Takekoshi, Masataka, Cheng, Xun-Jia, Nakata, Yuta, Takeuchi, Tsutomu, Ihara, Seiji
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 01-01-2004
• Subjects: Alkaline Phosphatase - genetics; Animals; Antibodies, Monoclonal - genetics; Antibodies, Protozoan - genetics; Antigens, Protozoan; Base Sequence; DNA, Recombinant - genetics; Dysentery, Amebic - diagnosis; Entamoeba histolytica; Entamoeba histolytica - immunology; Escherichia coli; Escherichia coli - genetics; Genetic Vectors; Humans; Immunoglobulin Fab Fragments - genetics; Microbial Immunology; PhoA gene; Recombinant Fusion Proteins - genetics
• ISSN: 1071-412X; 1098-6588
• DOI: 10.1128/CDLI.11.1.216-218.2004

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.