Description
This ELISA is intended for the quantitative determination of lactoferrin in stool. In a first incubation step, the lactoferrin in the samples is bound to polyclonal antibodies, immobilised to the surface of the microtiter wells. To remove all unbound substances, a washing step is carried out. In a second incubation step, a peroxidase-labelled conjugate (rabbit anti human lactoferrin) is added which recognises specifically the bound lactoferrin. After another washing step to remove all unbound substances, the solid phase is incubated with the substrate, tetramethyl-benzidine (TMB), which reacts with the peroxidase. An acidic stop solution is added to stop the reaction. The colour changes from blue to yellow. The intensity of the yellow colour is directly proportional to the concentration of lactoferrin. 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. Lactoferrin, present in the patient samples,is determined directly from this curve