Maturation of antibody response to nanogram quantities of A.fumigatus allergen
Proteins originated from different allergens penetrate mucosal surfaces at extremely low concentrations. However they are recognized by immune system and can induce IgE production leading to hypersensitivity to the allergen. The maturity of this response is not known. The purpose of this work was to estimate minimal doses of allergen recognized by the immune system and compare affinity of IgG and IgE antibodies induced by immunization with high or low doses of allergens. A major allergen Asp f 2 from A. fumigatus fungi was used to immunize BALB/c mice in the range from 100 ug to 1 pg per mouse. Asp f 2 was injected s.c. 10 times every day in PBS. Specific IgG was found in mice immunized with 50 ng/mouse and higher. Memory response was detected 2 months after immunization without drops in the antibody titer. Asp f 2 specific IgE was found at the dilution 1:20 in all groups immunized with 50 ng/mouse and higher. To estimate affinity of IgG and IgE antibodies produced during immunization with nanogram quantities of allergens we have compared dissociation constants (Kd) of antibodies produced in mice immunized with 500 ng of Asp f 2 (total dose) in PBS and in mice immunized with a standard single s.c. immunization of mice with 100 ug of Asp f 2 in CFA. Kd were estimated by inhibitory ELISA. Immunization of mice with high dose of Asp f 2 in CFA induced high affinity IgG (Kd=10 nM) 10 days after immunization. This affinity was the same when measured 2 months later. No IgE was found in this group. In mice immunized with nanogram quantities of Asp f 2 in PBS IgG had low affinity (Kd>10 uM) 7 days after the last injection, but it was increasing steadily during the next month (Kd=200; 100; and 30 nM at weeks 2, 4, and 5, accordingly). High affinity antibodies were still registered 2 months after immunization. The estimation of IgE affinity has shown that IgE antibodies maturated with the same rate. At week 2 the affinity of IgE was equal to the affinity of IgG (Kd=200 nM) and it increased during the month to the same level (Kd=45 nM) as of IgG. IgE production steadily subsided and no Asp f 2 specific IgE was found at week 7 after immunization. These results demonstrate that i) immune system can recognize effectively nanogram quantities of foreign antigens; ii) immunization with antigen in soluble form is extremely effective when it is injected several times to be displayed to the immune system; iii) immune response to nanogram quantities of Asp f 2 is long-lasting; iv) immunization with soluble antigen induces both IgG and IgE; v) the affinity of IgG and IgE increases steadily in mice immunized with soluble antigen in nanogram quantities while it increases sharply in mice immunized with high dose antigen in CFA; vi) however, finally the affinity of IgG is comparable between groups; vii) the affinity of IgG and IgE is the same.
- AAA 2nd (2006)
Full conference title:
2nd Advances against Aspergillosis