The whole blood concentration of tacrolimus is required for therapeutic drug monitoring of this immunosuppressive drug. Abnormal tacrolimus levels affect its efficacy or toxicity, leading to changes in its dosage. Here, we report analytical interference in the affinity column-mediated immunoassay tacrolimus method on the Xpand autoanalyzer in a kidney transplant human immunodeficiency virus-infected patient. Tacrolimus concentrations obtained by affinity column-mediated immunoassay are 3- to 7-fold higher than measurements with the enzyme multiplied immunoassay technique assay. The cause of this interference remains unknown. However, it would be necessary to identify this type of interference to measure tacrolimus concentration with another method to avoid analytical error, which may lead to a poor clinical outcome.