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Production of Nfa1-specific monoclonal antibodies that influences the in vitro cytotoxicity of Naegleria fowleri trophozoites on microglial cells.

Lee, YJ; Kim, JH; Jeong, SR; Song, KJ; Kim, K; Park, S; Park, MS; Shin, HJ
Parasitology research, 101(5):1191-1196, 2007
Journal Title
Parasitology research
Naegleria fowleri, agent of fatal primary amoebic meningoencephalitis, appears to induce cytotoxicity mechanically through its contact with the cell. The nfa1 gene cloned from a cDNA library of pathogenic N. fowleri by immunoscreening consists of 360 bp and expresses a 13.1-kDa recombinant protein (rNfa1) that demonstrated localization in the pseudopodia when examined using immunocytochemistry. To study the mechanisms involved in N. fowleri cytotoxicity, we developed a large volume of rNfa1-specific monoclonal antibody (McAb) against a 17-kDa His-tag fusion rNfa1 protein using a cell fusion technique. We established eight McAb-producing hybridoma cells. The antibodies were all immunoglobulin G2b and reacted strongly with a 17-kDa band representing the rNfa1 fusion protein in Western blotting, demonstrating immunoreactivity to the Nfa1 protein in pseudopodia (especially in the food cups) of N. fowleri trophozoites. A 51Cr-release assay indicated N. fowleri cytotoxicity by demonstrating that it eliminated 37.8, 60.6, and 98.8% of the target (microglial) cells 6, 12, and 24 h after co-incubation, respectively. When an anti-Nfa1 McAb was added to the coculture system, N. fowleri cytotoxicity decreased to 29.8, 44.1, and 66.3%, respectively.
MeSH terms
AnimalsAntibodies, Monoclonal/immunology*Antibodies, Protozoan/immunology*Antigens, Protozoan/geneticsAntigens, Protozoan/immunology*Cells, CulturedChromium Radioisotopes/metabolismMicroglia/parasitology*Naegleria fowleri/immunologyNaegleria fowleri/pathogenicity*Protozoan Proteins/geneticsProtozoan Proteins/immunology*RatsRecombinant Fusion Proteins/geneticsRecombinant Fusion Proteins/immunologyTrophozoites/immunology*
Appears in Collections:
Journal Papers > School of Medicine / Graduate School of Medicine > Pediatrics & Adolescent Medicine
Journal Papers > School of Medicine / Graduate School of Medicine > Microbiology
AJOU Authors
정, 석률송, 경주김, 경민박, 선박, 문성신, 호준
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