Albumin ELISA

This Enzyme-Linked Immunosorbent Assay (ELISA) is a two-step assay for the ultra-sensitive determination of human albumin in stool and urine. A polyclonal rabbit antibody specific for human albumin is immobilized on a microtiter plate and a second anti-albumin antibody is conjugated to peroxidase. In a first incubation step, the albumin in the samples is bound to the immobilized antialbumin antibodies. A washing step is carried out to remove all unbound substances. In a second incubation step, a peroxidase-labeled anti-albumin antibody is added. After another washing step, to remove all unbound substances, a peroxidase substrate, tetramethylbenzidine, is added. The enzymatic reaction is terminated by an acidic stop solution, whereby the color converts from blue to yellow. The intensity of the yellow color is directly proportional to the concentration of albumin in the sample. A dose response curve of the absorbance unit (optical density, OD) versus concentration is generated using the results obtained from the calibrators. Albumin in the samples is determined directly from this curve.