Construction and validation of Plasmodium antigens surface display library in mammalian cells.

Abstract

Malaria infection has been ongoing for the last 50,000 years, yet unlike other

organisms, vaccine is still under-development. Experimental immunization with

whole parasites live, physically or genetically attenuated provides successful

protection in mice and humans. In order to identify the antigens involved in efficient

immune response, we have developed tools and an assay to aid in the process of

identification. Malaria antigens from p.yoelii and p.falciparum were cloned and

expressed in mammalian cell surface display system. Using flow cytometry, nontransfected

and transfected cells were characterized by labeling with anti-myc and

the malaria fusion proteins were tested with anti-malaria antibodies. Similar

transfection rates were observed in the transfected HEK293 and HEK293T cells but

under drug pressure, only HEK293 cells could be used for positive transfectants

selection. Culturing of plasmid containing cells in 200μg/ml geneticin increased

percentage of cells containing plasmids. Transfected cells were specifically labeled

with anti-malaria antibodies, proving reliability of the assay for use in screening for

antigens recognized in immunized mice or humans.