Description
The assay utilises the “sandwich” technique with two selected antibodies that recognise human lysozyme. Standards, controls and diluted samples, which are assayed for human lysozyme, are added into the wells of a micro plate coated with a high affine anti-human lysozyme antibody. During the first incubation step, lysozyme is bound by the immobilised antibody. Then a peroxidase-conjugated anti-human lysozyme antibody is added into each microtiter well and a “sandwich” of capture antibody – human lysozyme – peroxidase-conjugate is formed. Tetramethylbenzidine is used as peroxidase substrate. Finally, an acidic stop solution is added to terminate the enzymatic reaction. The colour changes from blue to yellow. The intensity of the yellow colour is directly proportional to the concentration of lysozyme. A dose response curve of the absorbance unit (optical density, OD at 450 nm) vs. concentration is generated using the values obtained from the standards. Lysozyme, present in the samples, is determined directly from this curve