Histamine, ELISA

In the first part of the procedure, histamine is quantitatively acylated to N-acyl histamine. The subsequent competitive ELISA uses the microtiter plate format. The antigen is bound to the solid phase of the microtiter plate. The acylated standards, controls and samples compete with the solid phase bound analytes for a fixed number of antibody binding sites. After the system is in equilibrium, free antigen and free antigen-antibody complexes are removed by washing. The antibody bound to the solid phase is detected by an anti-goat IgG-peroxidase conjugate using TMB as a substrate resulting in a colour reaction. The reaction is monitored at a wavelength of 450 nm. Quantification of unknown samples is achieved by comparing their absorbance with a reference curve prepared with known standard concentrations