Anna Becker1, Janne T Backman2, Outi Itkonen1. 1. HUSLAB, University of Helsinki and Helsinki University Hospital, Helsinki, Finland. 2. Department of Clinical Pharmacology, University of Helsinki and Helsinki University Hospital, Helsinki, Finland; Individualized Drug Therapy Research Program, Faculty of Medicine, University of Helsinki, Helsinki, Finland.
Abstract
INTRODUCTION: Life-long monitoring of immunosuppressive drugs (ISDs) in blood is essential after organ transplantation. However, the ISD concentrations vary depending on the assay employed. ISDs are strongly bound to cytoplasmic proteins in erythrocytes in circulation. Therefore, the relatively rapid sedimentation of blood cells in whole blood samples may affect the results when using liquid handling robots. METHODS: We used 1115 blood samples from outpatients and ward patients with kidney (n = 373), liver (n = 101), heart (n = 29) and bone marrow (n = 155) transplant. Whole blood samples were pretreated by protein precipitation. Alternatively, the samples were hemolyzed by freezing prior to precipitation. ISDs were analyzed by a 2-plexing liquid chromatography tandem mass spectrometry (LC-MS/MS) assay and commercial chemiluminescent microparticle immunoassays (CMIA). RESULTS: The difference between the two sample preparation practices was negligible (<2%). Overall, the measured ISD concentrations in patient samples were lower by LC-MS/MS than by CMIA. The difference was the largest (20.2%) and the smallest (9.1%) in samples from liver and from heart transplant patients, respectively. CONCLUSIONS: CMIA overestimates blood ISD concentrations as compared to LC-MS/MS. The extent of the difference was found to be organ transplant dependent. The ISDs can be quantitatedeither from intact or hemolyzed blood samples.
INTRODUCTION: Life-long monitoring of immunosuppressive drugs (ISDs) in blood is essential after organ transplantation. However, the ISD concentrations vary depending on the assay employed. ISDs are strongly bound to cytoplasmic proteins in erythrocytes in circulation. Therefore, the relatively rapid sedimentation of blood cells in whole blood samples may affect the results when using liquid handling robots. METHODS: We used 1115 blood samples from outpatients and ward patients with kidney (n = 373), liver (n = 101), heart (n = 29) and bone marrow (n = 155) transplant. Whole blood samples were pretreated by protein precipitation. Alternatively, the samples were hemolyzed by freezing prior to precipitation. ISDs were analyzed by a 2-plexing liquid chromatography tandem mass spectrometry (LC-MS/MS) assay and commercial chemiluminescent microparticle immunoassays (CMIA). RESULTS: The difference between the two sample preparation practices was negligible (<2%). Overall, the measured ISD concentrations in patient samples were lower by LC-MS/MS than by CMIA. The difference was the largest (20.2%) and the smallest (9.1%) in samples from liver and from heart transplant patients, respectively. CONCLUSIONS: CMIA overestimates blood ISD concentrations as compared to LC-MS/MS. The extent of the difference was found to be organ transplant dependent. The ISDs can be quantitatedeither from intact or hemolyzed blood samples.