Preparation And Identification Of Two Monoclonal Antibodies

Purpose Candida, which cause candidiasis, has became a commensal pathogen of nosocomial infection. And Candiada albicans(C.albicans), as one of the major pathogens agents of candidiasis,invases the hosts through dimorphic transition, attachment and then colonization on surface of endothelial cells and mucous membrane, and so on. C. albicans not only can cause the lesions of skin and mucous membrane, but also can cause the infection of viscera.It is necessary to identify this species rapidly and early in diagnosis of systemic candidiasis, which can be of benefit in selecting the most effective therapies. But untill now, the method, particularly with the recent sensitive and specific identification of early infection, is outdated. At present, the clinical studies has focused on detection of the specific antigens and metabolites of C.albicans. Two strains of lymphocyte hybridom were prepared,which provide two new laboratory methods for early and rapid diagnosis of systemic Candidasis. At the same time study primarly the function of McAb against C.albicans Germ Tubes, which inhibit dimorphic transition in vitro.Methods Two Monoclonal antibodies separately against strain ATCC-90028 of C.albicans germ tube and Yeast enoalse were prepared and identified by the lymphocyte hybridom technique, through immuning BALB/c mice using the antigen of enolase of yeast and the antigen of blastocondida of standard strain ATCC-90028 with germ tube.The results of germ tubes induction, which were induced by NCS , and with the participation of McAb against C. albicans Germ Tubes, were observed, and the results of this experimentation were dealed with statistics.Resultes Two strains lymphocyte hybridom named Mab03.3G5-F3-F5,Mab 03.4G6-F3-C7, which created McAb against enolase of yeast ,were prepared by lymphocyte hybridom technique, and Mab03.3G5-F3-F5 was identified. The titer of McAb of Mice hydroperitonia reached to 1:12800;The Mab03.3G5-F3-F5 belonged to IgG1 Kappa isotype. Result from Western Blotting manifested that the McAb could specificlly reacted to the antigen of enolase (48kD) of Yeast . It is the McAbs against enolase of yeast, which was proved by SDS-PAGE and Western Blotting.Only one strain lymphocyte hybridom named Mab03.2C1-C2 creating McAb against specific antigen of C. albicans germ tube, was prepared by lymphocyte hybridom technique. The lymphocyte hybridom of Mab03.2C1-C2 was screened by IIF, and fluorence was observed on the surface of germ tubes but not on blastoconidia. The titer of McAb reached to 1:25600; The McAb belonged to IgG1 Lambda isotype. McAb Mab03.2C1-C2 reacting exclusively to C.albicans germ tubes was studied by Western Blotting. It was the McAb against specific antigen of C. albicans germ tubes of ATCC-90028, which was proved by SDS-PAGE and Western Blotting.It was showed that McAb Mab03.2C1-C2 could reacted with component of 156kD within cell wall of C.albicans germ tubes extract. We found that the induction of C. albicans germ tubes were inhibited by Mab- 03.2C1-C2, from what was observed in the experiment with McAb against specific antigen of C. albicans germ tubes.Conclusions One strain lymphocyte hybridom against specific antigen of C. albicans germ tubes and Five strains lymphocyte hybridom against enolase of yeast were prepared and identified in this study, and conclusion can be made that Mab03.2C1-C2 identify component 156kD within cell wall of C.albicans germ tubes extract. This is a basal study of the pathogenic experiment of protection during systemic C.albicans using the Mab03.2C1-C2, which forms the foundation for next investigation for protective mechanism of McAb in anti-C.albicans ,and using both strain McAbs to develop diagnose reagent.