*4.5. Procedure and Judgment of the ICA*

When using the prepared ICA to detect ZEN, the ZEN standard solution or sample extraction solution (100 µL) was put onto the sample pad of the ICA strip. The ICA detection could be finished after 5 min under the capillary action and immune reaction. At that moment, the detection results of the red color intensity would be reflected on the T line and C line, which were inversely proportional with the ZEN levels in the detection solutions. The depth of the red color intensity could be evaluated by two models—the naked eye and strip reader—as shown in Figure 1. Using model I, the ICA results could be judged as the following four cases: (1) negative (−), the ZEN was less than the LOD or free; (2) weakly positive (±), the ZEN was more than the LOD but lower than the concentration of the complete inhibition of the immune reaction; (3) positive (+), the ZEN was above the concentration of the complete inhibition of the immune reaction; and (4) ineffective, the ICA strip was not working. Using model II, the ICA results could be detected digitally. The gray values of the T lines could be obtained, and the inhibition ratios could be used to establish a linear relationship with the concentrations of ZEN levels using model II.
