RESUMEN
We evaluated a new protocol for measurement of cyclosporine A (CsA) 2 H after dose (C2) on the V-Twin® analyzer. Imprecision, recovery, and linearity were determined using CsA-spiked blood pools. Accuracy was evaluated using specimens from renal, cardiac, and liver transplant patients, and results were compared with those from liquid chromatography-tandem mass spectrometry (LC-MS/MS) and the Abbott TDx®/TDxFLx® assay. Cross-reactivity and interferences were assessed in the presence of 800 ng/mL CsA. Imprecision coefficients of variation were 3.3%-4.8% (within run) and 5.9%-8.7% (total). Recovery was within 10% of the expected values. Linearity was 350-2,000 ng/mL. Calibration was stable for ≥ 2 weeks. Method comparison showed regression statistics: V-Twin® = 1.01 × LC tandem MS + 36.1, r = 0.971; V-Twin® = 1.13 × Abbott - 92.4, r = 0.969. Metabolite cross-reactivity and interference (endogenous substances and drugs) were within ±10%. The C2 protocol on the V-Twin® analyzer provides acceptable assay performance and accurate determination of whole blood CsA drawn at 2 H after dose.
Asunto(s)
Ciclosporina/sangre , Técnicas para Inmunoenzimas/instrumentación , Inmunosupresores/sangre , Calibración , Reacciones Cruzadas , Ciclosporina/inmunología , Trasplante de Corazón , Humanos , Técnicas para Inmunoenzimas/métodos , Trasplante de Riñón , Trasplante de Hígado , Sensibilidad y EspecificidadRESUMEN
Buprenorphine is now increasingly prescribed as an alternative to methadone for the treatment of heroin addiction. Because of its potency (dosage usages from 0.2 mg to 8 mg), the drug concentrations in body fluids are normally very low. Here, we report the first recombinant glucose-6-phosphate dehydrogenase (G6PDH)-based homogeneous immunoassay (EMIT-type assay) for free buprenorphine and free norbuprenorphine in urine. The antibody used in this assay cross-reacts nearly identically with buprenorphine and norbuprenorphine and, at the same time, has less than 1% cross-reactivity with a wide range of commonly prescribed opiates, particularly those structurally related compounds such as morphine, codeine, and dihydrocodeine. More importantly, this assay has a low detection limit of 1 ng/mL for buprenorphine or norbuprenorphine. Further evaluation of this technique using gas chromatography-mass spectrometry (GC-MS) of authentic urine samples demonstrated that the accuracy of the assay is greater than 95%. Because this assay is designed to measure the free drugs in urine, it resulted in simplification for GC-MS or liquid chromatography-MS confirmation methods that did not require urine hydrolysis before solid-phase or liquid-liquid extraction.
Asunto(s)
Analgésicos Opioides/orina , Buprenorfina/análogos & derivados , Buprenorfina/orina , Glucosafosfato Deshidrogenasa , Analgésicos Opioides/uso terapéutico , Buprenorfina/uso terapéutico , Reacciones Cruzadas , Técnica de Inmunoensayo de Enzimas Multiplicadas , Cromatografía de Gases y Espectrometría de Masas/métodos , Dependencia de Heroína/tratamiento farmacológico , Humanos , Proteínas Recombinantes , Reproducibilidad de los Resultados , Detección de Abuso de Sustancias/métodosRESUMEN
New application protocols for the Emit(®) II Plus 6-Acetylmorphine Assay for human urine screening have been developed on the ADVIA(®) 1800 and 2400 Chemistry Systems. Precision was evaluated at the cutoff and ±25% controls. Recovery and linearity were studied by spiking 6-acetylmorphine (6-AM) into human urine pools. Method comparison was evaluated using urine specimens and the results were compared to those obtained from the predicate Analyzer (V-Twin(®)). Cross-reactivity with structurally related drugs was assessed at high cross-reactant concentrations. Potential interferences were assessed in the presence of 7.5 and 12.5 ng/mL of 6-AM. The qualitative repeatability coefficients of variation (CV's) ranged from 0.40 to 0.90% and the within-lab CV's ranged from 1.3 to 3.5%. In analyte units (ng/mL), the repeatability CV's ranged from 1.9 to 4.3% and the within-lab CV's ranged from 3.7 to 6.1%. The limit of detection of the assay was found to be 2.5 ng/mL on both instruments. Recovery was within 20% of expected value. Linearity was 2.5-20 ng/mL. Method comparison showed 100% agreement with the predicate analyzer. The assay had minimal cross-reactivity to structurally related opioids including with morphine, morphine-3-glucuronide, morphine-6-glucuronide. No interference was observed with endogenous interferences.
Asunto(s)
Técnica de Inmunoensayo de Enzimas Multiplicadas/instrumentación , Dependencia de Heroína/diagnóstico , Derivados de la Morfina/orina , Narcóticos/orina , Detección de Abuso de Sustancias/instrumentación , Toxicología Forense/instrumentación , Humanos , Límite de Detección , Reproducibilidad de los ResultadosRESUMEN
We evaluated the performance of Emit(®) II Plus 6-Acetylmorphine Assay for human urine screening on the Viva-E(®) analyzer. Precision was evaluated using the cutoff and ±25% controls. Recovery and linearity were studied by spiking 6-acetylmorphine (6-AM) into human urine pools. Accuracy was evaluated using urine specimens and the results were compared to those from GC/MS. Cross-reactivity with structurally related drugs was assessed at different cross-reactant concentrations. Interferences were assessed in the presence of 7.5 and 12.5 ng/mL of 6-AM. The qualitative repeatability coefficients of variation (CV's) ranged from 0.3% to 0.4% and the within-lab CV's ranged from 2.0% to 2.2%. In analyte units (ng/mL), the repeatability CV's ranged from 1.3% to 2.2% and the within-lab CV's ranged from 2.6% to 4.3%. The limit of detection of the assay was found to be 2.1 ng/mL. Recovery was within 15% of expected value. Linearity was 2.1-20 ng/mL. Method comparison showed 99% agreement with GC/MS. The assay had minimal cross-reactivity with morphine, morphine-3-glucuronide, morphine-6-glucuronide and other opioids. No interference was observed with endogenous interferences and structurally unrelated drugs. The assay correctly classified CAP survey samples. The Emit(®) II Plus 6-Acetylmorphine Assay will be a suitable screening method for urine specimens in both qualitative and semi-quantitative analyses.