Sirolimus is an immunosuppressive agent administered as prophylaxis of acute rejection to patients after kidney transplantation. Therapeutic drug monitoring (TDM) of whole blood is an important part of immunosuppressive therapy. At present, two methods of measuring drug concentrations are available: the reference method is high-performance liquid chromatography (HPLC) with ultraviolet (UV) or mass spectrometry (MS) detection and a second technique is the IMx sirolimus assay, which is an enzyme immunoassay using microparticles coated with anti-sirolimus antibodies (MEIA). The objective of this study was to compare the two methods. We examined a group of 42 patients receiving sirolimus after kidney transplantation. Blood was taken during routine ambulatory visits. The drug concentration in blood was performed at the same time by the two methods. To compare the methods, a statistical analysis was performed, yielding: r-value = 0.939 (r(2)); slope = 1.04; intercept = +0.38. The mean concentration of sirolimus was higher in the immunoassay than in the HPLC method namely, 9.7 +/- 6.4 ng/mL versus 8.9 +/- 5.8 ng/mL respectively. The HPLC method showed high sensitivity and specificity, but it was time consuming and labor intensive. The MEIA method is burdened with a high risk of methodologic error, due to its lack of specificity caused by cross-reactions with drug metabolites. We concluded that HPLC with its high sensitivity and analytical specificity is still the reference method; however, MEIA may be a fast method for use in clinical practice.