Performance of a no-pretreatment tacrolimus assay on the Dade Behring Dimension RxL clinical chemistry analyzer.

BACKGROUND: Therapeutic drug monitoring for tacrolimus is important for organ transplant patients receiving this immunosuppressant. Current available assays for tacrolimus require sample pre-treatment and operate in a batch mode. Here a no-pretreatment tacrolimus assay, performed on the Dade Behring Dimension analyzer is compared to the Abbott IMx tacrolimus assay and to an LC/MS/MS method. METHODS: Whole blood samples from 2 medical centers and different transplant types (kidney n=103, liver n=81, heart n=27, pancreas n=16, bone marrow n=9, [corrected] lung n=7), were obtained and tacrolimus quantified by each of the 3 assays. RESULTS: The lower limit of the linear range was 1.2 ng/ml on the Dimension assay. Total imprecision was 9.8% and within-run imprecision was 9.6% at a tacrolimus concentration of 3.4 ng/dL. Passing-Bablock regression analysis determined the following relationships: DIMN=(1.16) LC/MS - 0.43, r=0.90 and DIMN=(0.99)IMx - 0.35, r=0.87. CONCLUSIONS: The Dade Behring Dimension [corrected] Tacrolimus assay has adequate imprecision and correlates well with the reference method of LC/MS/MS. The assay appears suitable for clinical use, and has the advantages of not requiring a pretreatment step and the ability to be performed in a random-access mode.

Successive neuron loss in the thalamus and cortex in a mouse model of infantile neuronal ceroid lipofuscinosis.

Infantile neuronal ceroid lipofuscinosis (INCL) is caused by deficiency of the lysosomal enzyme, palmitoyl protein thioesterase 1 (PPT1). We have investigated the onset and progression of pathological changes in Ppt1 deficient mice (Ppt1-/-) and the development of their seizure phenotype. Surprisingly, cortical atrophy and neuron loss occurred only late in disease progression but were preceded by localized astrocytosis within individual thalamic nuclei and the progressive loss of thalamic neurons that relay different sensory modalities to the cortex. This thalamic neuron loss occurred first within the visual system and only subsequently in auditory and somatosensory relay nuclei or the inhibitory reticular thalamic nucleus. The loss of granule neurons and GABAergic interneurons followed in each corresponding cortical region, before the onset of seizure activity. These findings provide novel evidence for successive neuron loss within the thalamus and cortex in Ppt1-/- mice, revealing the thalamus as an important early focus of INCL pathogenesis.

CNS-directed AAV2-mediated gene therapy ameliorates functional deficits in a murine model of infantile neuronal ceroid lipofuscinosis.

The neuronal ceroid lipofuscinoses (Batten disease) are a group of inherited neurodegenerative diseases characterized by the progressive intralysosomal accumulation of autofluorescent material in many cells, visual defects, seizures, cognitive deficits, and premature death. Infantile neuronal ceroid lipofuscinosis (INCL) has the earliest onset ( approximately 1.5 years of age) and is caused by a deficiency in the lysosomal enzyme palmitoyl protein thioesterase-1 (PPT1). Currently there is no effective treatment for children with INCL. In this study, newborn PPT1-deficient mice received two (cortex), four (cortex and hippocampus), or six (cortex, hippocampus, and cerebellum) bilateral intracranial injections of AAV2-PPT1. The AAV-treated animals had localized increases in PPT1 activity, decreased autofluorescent material, improved histologic parameters, and increased brain mass. In addition, the treated animals had dose-dependent improvements in a battery of behavioral tests and improved interictal electroencephalographic tracings. However, there was neither a significant decrease in seizure frequency nor an increase in longevity even in INCL animals receiving six injections. These data suggest that early treatment of INCL using gene transfer techniques can be efficacious. However, higher levels or a broader distribution of PPT1 expression, or both, will be required for more complete correction of this neurodegenerative disease.