Antibody testing in serum
According to the requirements of the reagent instructions, the
antibodies in the serum were detected by Enzyme-linked immunosorbent
assay(ELISA). In brief, 10 μg/ml Echinococcus multilocularis crude
antigen was coated on the ELISA plate and placed at 4°C overnight. The
plates were washed five times with PBST (0.05% Tween 20 PBS) and
blocked with 5% skim milk powder in PBST at 37°C for 1 h. After washing
five times with PBST, the plates were incubated with mouse serum (1:500)
in 5% skim milk powder in PBST for 2 h and washed five times with PBST
for 3 min. And one hundred microliters each of horseradish peroxidase
(HRP)‐conjugated anti‐mouse IgM, IgG, IgA(abcam, USA) and
IgE(Invitrogen, USA) were added enzyme plates and incubated at 37°C for
1 h. After washing, 100μl TMB Single-Component Substrate
solution(Solarbio, China) was added for 8–10 min, and the reaction was
stopped by 2 M H2SO4. The absorbance was measured at 450 nm using an
ELISA reader (Thermo Fisher, USA).