Immunoassays for low concentrations of albumin in urine

We have developed and validated simple, rapid immunoassays to measure concentrations of albumin in urine ranging from 5 to 200 mg/L. We use antiserum to human albumin, raised in sheep, and we separate the antibody-bound and free fractions of albumin by using a second antiserum, produced in donkeys, against the Fc fragment of sheep IgG. These two antisera can be mixed and added to assay tubes as a single reagent without inhibiting antigen binding. Samples or standards are incubated for at least 30 min with fluorescein-labeled albumin and the premixed antiserum reagent. After brief centrifugation, the supernate is discarded and the precipitate containing the bound fraction is dissolved and its fluorescence measured. Alternatively, 125I-labeled albumin can be used as tracer with all other reagents unchanged; in this case one simply counts the radioactivity of the bound fraction. A reference interval of less than 0.2-2.8 mg/mmol of creatinine was obtained for untimed, daytime, midstream urine specimens from 80 healthy subjects.