Caracterização parcial de fator(es) hemolítico(s) de Arthrographis kalrae

AUTOR(ES)
DATA DE PUBLICAÇÃO

2009

RESUMO

In this study, soluble components of Arthrographis kalrae have been investigated for immunogenicity/antigenicity and hemolytic activity. For immunogenicity/antigenicity assays, A. kalrae cell-free antigens (CFA), sonicated antigens (SA) and rabbit anti-CFA by immunodiffusion (ID)/enzyme linked immunosorbent assay (ELISA)/western blotting (WB) were used in addition to Sephadex G-100 and G-200 chromatography fractions analysis by ELISA. The hemolytic activity was performed with either sheep red blood cells (SRBC) or mouse red blood cells (MRBC) and mouse anti-CFA and mouse anti- MRBC sensitized with CFA (anti-MRBC-CFA). For mouse, CFA in its native form, previously heated CFA (56 ºC, 30 min) (hCFA) or treated with anti-MRBC-CFA and CFA Sephadex G-200 chromatography fractions were analyzed by hemolytic assay. Additionally, the reactivity of the anti-MRBC-CFA or anti-CFA with CFA Ags was analyzed by ELISA using Sephadex G-200 chromatography fractions or by WB. The results using rabbit serum demonstrated one peak of reactivity in fractions of Sephadex G-100 and two peaks of reactivity in Sephadex G-200 for CFA antigens. Results of WB showed that rabbit anti-CFA recognized several bands with distinct MM, being the profile differentiated in respect to the antigen preparation. Analysis of hemolytic activity resulted in lyses of the SRBC. Using MRBC, hemolytic activity was observed with CFA and hCFA and hemolytic activity decreased with CFA anti-MRBC-CFA treated samples, suggesting specificity of the antibodies to the factors responsible for hemolysis. Hemolytic activity was observed in chromatography G-200 fractions, also positive with anti-MRBC-CFA by ELISA. WB resulted in two main bands with MM ~252 kDa and ~79 kDa recognized by anti-MRBC-CFA. In conclusion, A. kalrae releases at least two main components with antigenic, immunogenic and with hemolytic activity, which are thermostable with MM >200kDa and ~79 kDa.

ASSUNTO(S)

enzime immunoassay microbiologia microorganismos patogênicos teste imunoenzimático microbiology pathogenic microorganisms

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