Fig. 4

Optimization of indirect-ELISA by checkerboard titration. Positive bar: OD₄₅₀ value of H. contortus infected goat sera; Negative bar: OD₄₅₀ value of helminths frees goat sera; P/N: OD₄₅₀ value of positive/ OD₄₅₀ value of negative that showed on each bar and horizontal red dotted line is the cutoff line at 2.1 (right Y axis). P/N value more than 2.1 was determined to be positive while P/N value less than 2.1 was determined to be negative. The best condition (the highest P/N value) of each other was pointed by red arrows (↑). Graph a: Selection of antigen incubating condition; Graph b: Selection of blocking buffer; Graph c: Selection of optimal dilution of rabbit anti-goat IgG; Graph d: Selection of reactive time of serum; Graph e: Selection of blocking time; Graph f: Selection of reactive time of TMB. The optimum antigen incubating condition (37 °C 1 h and overnight at 4 °C), best blocking buffer (5% BSA), optimal dilution of rabbit anti-goat IgG (1:4000), best reactive time of serum (120 min), blocking (60 min) and reactive time of TMB (10 min) were observed