An HPLC/MS/MS assay for tacrolimus in patient blood samples. Correlation with results of an ELISA assay
Abstract
An HPLC/MS/MS assay for tacrolimus in whole blood using FR900520 as an internal standard was validated over the standard curve range of 0.100-10.040 ng ml-1. The calibration curve for tacrolimus in human blood gave a slope of 0.2481, an intercept of 0.007, and a correlation coefficient (r) of 0.9996, with no interference noted from human blood, analyte, or internal standard stock solutions. Use of EDTA or heparin as the preservative in blood resulted in no significant differences. Samples were stable for at least the time required to assay the maximum number of samples that could be placed in the automated system. The limit of sensitivity of the assay was set at the concentration of the lowest nonzero standard tested, i.e., 0.100 ng ml-1. However, validation of the assay to a limit of 0.010 ng ml-1 is currently underway. The within-run and between-run precision and accuracy of the method were determined for four quality control samples. The highest CV was seen at 0.1 ng ml-1 (17.6% within-run and 15.9% between-run), with other CV < 5%. The recovery ranged 79.6-81.3% for tacrolimus over the range 0.3-8.0 ng ml-1 and was 63.10 +/- 1.37% for FR900520. There was a linear correlation (r2 = 0.963) between assay results by HPLC/MS/MS ...Continue Reading
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