A highly sensitive enzyme-linked immunosorbent assay for the determination of tacrolimus in atopic dermatitis patients

A highly sensitive enzyme-linked immunosorbent assay method has been developed for the determination of tacrolimus in human blood samples. The assay is a modification of the previously published assay with improved sensitivity. Following extraction of tacrolimus with methanol and sulfosalicylic acid, the samples are incubated for 2 h at room temperature on a Nunc Maxisorb plate that has been treated for nonspecific binding by precoating with polyclonal antibody. The analysis of human blood following the standard addition of tacrolimus (0.02-5.0 ng/ml) demonstrated excellent precision and accuracy over a 6-day period. The interday and intraday co-efficients of variation were 6.0-28.9 and 3.9-15.2%, respectively. The limit of quantitation was 0.05 ng/ml. The method was used to quantitate blood concentration of tacrolimus in patients following the administration of tacrolimus ointment.