Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 19 de 19
Filter
1.
Liver Transpl ; 14(12): 1745-51, 2008 Dec.
Article in English | MEDLINE | ID: mdl-19025918

ABSTRACT

Mycophenolic acid (MPA) is used to prevent graft rejection. The methods used for determining the plasma MPA concentration in liver transplant recipients are the enzyme-multiplied immunoassay technique (EMIT), high-performance liquid chromatography with ultraviolet detection (HPLC-UV), and most recently mass spectrometry. EMIT has been reported to overestimate the MPA concentration by 30% to 35% in comparison with HPLC-UV. Recently, a new automated enzymatic assay based on inosine monophosphate dehydrogenase inhibition has been designed. The aim of the present investigation was to compare this technique with validated HPLC-UV in adult liver transplant recipients treated with tacrolimus or cyclosporine. One hundred seventy-six samples from 50 adult liver transplant recipients were analyzed with both techniques. Patients received mycophenolate mofetil (2 or 3 times daily) coadministered with cyclosporine microemulsion (n = 18) or tacrolimus (n = 32). Samples were drawn over an interdose interval during the early or late posttransplantation period. The Passing-Bablok regression and Bland-Altman plot were used to compare the 2 techniques. The Passing-Bablock regression, calculated from 166 samples, showed very good agreement between the enzymatic assay and the HPLC-UV method: enzymatic assay = 1.0204 (95% confidence interval, 0.9942, 1.0478) x HPLC-UV + 0.0201 (-0.0442, 0.0882). No significant bias was found between the techniques (Bland-Altman plot), and the median relative difference was 2.7% (95% confidence interval, -0.4, 6.6). In conclusion, the enzymatic assay showed an excellent correlation with HPLC-UV. Therefore, this method was proved valid and reliable for the monitoring of the plasma MPA concentration in adult liver transplant recipients treated with cyclosporine microemulsion or tacrolimus.


Subject(s)
Drug Monitoring/methods , Enzyme Multiplied Immunoassay Technique , Immunosuppressive Agents/blood , Liver Transplantation , Liver/metabolism , Mycophenolic Acid/blood , Adult , Chromatography, High Pressure Liquid/methods , Drug Monitoring/standards , Enzyme Multiplied Immunoassay Technique/standards , Female , Graft Rejection/prevention & control , Humans , Immunosuppressive Agents/administration & dosage , Immunosuppressive Agents/pharmacokinetics , Male , Middle Aged , Mycophenolic Acid/administration & dosage , Mycophenolic Acid/pharmacokinetics , Ultraviolet Rays
2.
Ther Drug Monit ; 26(6): 609-19, 2004 Dec.
Article in English | MEDLINE | ID: mdl-15570184

ABSTRACT

Mycophenolic acid (MPA) is an immunosuppressive drug partly metabolized to MPA-glucuronide (MPAG), which is pharmacologically inactive. The currently available enzyme-multiplied immunoassay technique (EMIT) has been reported to overestimate MPA plasma concentration in clinical samples when compared with HPLC techniques. The aims of this study were to design and validate a specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) technique for the determination of MPA and MPAG using a low plasma volume and a simple sample preparation procedure; then to compare it with EMIT for the determination of MPA in plasma samples collected over an interdose interval at different posttransplantation periods (days 3, 7, and 30 and after 3 months) in 25 renal transplant recipients orally administered cyclosporine and mycophenolate mofetil twice daily, to investigate the origins of the differences between techniques. The LC-MS/MS technique developed showed limits of quantification (LOQs) of 0.1 mg/L and 1 mg/L for MPA and MPAG, respectively, and was linear, accurate, and precise from these LOQs up to 30 mg/L for MPA and 300 mg/L for MPAG. EMIT gave similar results to LC-MS/MS for spiked quality control samples (in a synthetic matrix or in drug-free plasma) but significantly overestimated MPA levels in clinical samples: EMIT - LC-MS/MS = +61.39% +/- 57.94%, with large variations depending on patients, time elapsed since transplantation, sampling time, and concentration levels. These results confirmed the known overestimation of the EMIT assay compared with a specific method and showed that the magnitude of this overestimation depended on sampling time and time after transplantation.


Subject(s)
Drug Monitoring/methods , Enzyme Multiplied Immunoassay Technique , Kidney Transplantation , Mycophenolic Acid/blood , Analysis of Variance , Chromatography, High Pressure Liquid/methods , Enzyme Multiplied Immunoassay Technique/standards , Gas Chromatography-Mass Spectrometry/methods , Graft Rejection/blood , Graft Rejection/drug therapy , Humans , Least-Squares Analysis , Mycophenolic Acid/therapeutic use , Statistics, Nonparametric
3.
Ann Clin Biochem ; 40(Pt 6): 697-700, 2003 Nov.
Article in English | MEDLINE | ID: mdl-14629812

ABSTRACT

BACKGROUND: The Dade Behring Syva(R) EMIT (enzyme multiplied immunoassay technique) method for the measurement of tacrolimus in whole blood was evaluated against the Abbott IMx(R) microparticle enzyme immunoassay (MEIA) method. EMIT measures tacrolimus colorimetrically, whereas MEIA measures the analyte using fluorimetry. Both methods incorporate a protein precipitation step prior to measurement. METHOD: Whole blood specimens were treated by two types of precipitation technique followed by analysis for tacrolimus by either MEIA or EMIT on the Bayer Advia 1650. Linearity and precision were assessed and correlation analysis performed to evaluate the EMIT assay on the Bayer Advia 1650. RESULTS: The EMIT tacrolimus assay was linear over the concentration range 0.0-22.0 micro g/L; the limit of detection was 1.2 micro g/L. Correlation between the Syva EMIT and IMx tacrolimus assays was excellent (r = 0.959) and no significant bias existed between the two methods (mean difference, delta = 0.221 micro g/L). Calibration data for the EMIT assay was stable for a period of 24-48 h on the Advia between runs. CONCLUSION: The Syva EMIT assay for the measurement of tacrolimus in whole blood is suited for daily routine use on the Bayer Advia 1650.


Subject(s)
Enzyme Multiplied Immunoassay Technique/instrumentation , Enzyme Multiplied Immunoassay Technique/standards , Tacrolimus/blood , Blood Chemical Analysis , Calibration , Tacrolimus/immunology
4.
Clin Biochem ; 36(4): 313-6, 2003 Jun.
Article in English | MEDLINE | ID: mdl-12810161

ABSTRACT

OBJECTIVES: Evaluate the performance of the new pretreatment (NPT) reagent for use with the EMIT cyclosporine A (CsA) assay. DESIGN AND METHODS: Samples from transplant patients receiving CsA were tested using a COBAS MIRA S. RESULTS: A downward shift in target values for commercial controls was observed using the NPT reagent. There was excellent correlation for patient results when comparing the NPT reagent with methanol pretreatment. CONCLUSION: The NPT reagent is an acceptable substitute for methanol in the EMIT(R) extraction procedure for CsA.


Subject(s)
Cyclosporine/blood , Enzyme Multiplied Immunoassay Technique/standards , Reagent Kits, Diagnostic/standards , Enzyme Multiplied Immunoassay Technique/instrumentation , Humans , Reproducibility of Results
6.
Ther Drug Monit ; 21(2): 256-8, 1999 Apr.
Article in English | MEDLINE | ID: mdl-10217349

ABSTRACT

A modified EMIT 2000 digoxin assay was developed on the Cobas Mira plus analyzer for the determination of very low plasma concentrations of the drug. The major modifications were a higher plasma volume withdrawn during the analysis step and calibration curves constructed in the range 0-2 ng/ml using calibrators made up with biological matrix. Assays were controlled with an internal, four-level quality control (targets: 0.15; 0.60; 1.70; 2.70 ng/mL). The within-day and day-to-day mean observed values +/- SD (n = 10) of these quality controls were 0.14 +/- 0.02 and 0.15 +/- 0.02 ng/mL; 0.57 +/- 0.01 and 0.64 +/- 0.03 ng/mL; 1.55 +/- 0.06 and 1.62 +/- 0.04 ng/mL, 2.82 +/- 0.09 and 2.82 +/- 0.12 ng/mL, respectively. The detection and the quantification limits were 0.02 and 0.08 ng/mL, respectively. No significant difference was observed between digoxin plasma concentrations measured by the original and the modified EMIT 2000 digoxin assay in 25 plasma specimens, ranging from 0.4 to 3.0 ng/mL, from patients receiving the drug. This modified digoxin EMIT 2000 assay was subsequently used to study digoxin pharmacokinetics after each of 18 healthy volunteers was administered a single 0.5 mg oral dose. The pharmacokinetic parameters found in this study were in accordance with the literature in healthy subjects, using radioimmunoassay (RIA) for digoxin plasma concentration determinations. In conclusion, the lower limit of quantification of this modified EMIT 2000 digoxin assay is similar to that of RIA and can serve as a valuable screen for digoxin pharmacokinetic interactions studies.


Subject(s)
Cardiotonic Agents/blood , Cardiotonic Agents/pharmacokinetics , Digoxin/blood , Digoxin/pharmacokinetics , Drug Monitoring/standards , Enzyme Multiplied Immunoassay Technique/standards , Humans , Quality Control , Reference Values , Sensitivity and Specificity
7.
Ther Drug Monit ; 20(2): 202-8, 1998 Apr.
Article in English | MEDLINE | ID: mdl-9558135

ABSTRACT

The authors evaluated the performance characteristics of the Abbott AxSYM Vancomycin II immunoassay in sera of patients with (n = 93 samples) and without (n = 327 patients) renal dysfunction. Correlation of vancomycin measurements with the Abbott AxSYM Vancomycin, Abbott TDx/TDxFLx, Syva enzyme-multiplied immunoassay technique (EMIT), DuPont automated chemistry analyzer (ACA), and high-performance liquid chromatography methods showed acceptable correlation as indicated by: slope values >0.95, r-values >0.97, y-intercepts <1.7 microg/ml, and S(y/x) ranging from 9% to 15% of the average vancomycin value. The AxSYM Vancomycin II assay showed acceptable correlation with AxSYM vancomycin, TDx/TDxFLx, and high-performance liquid chromatography methods in 93 samples from patients with renal dysfunction. This monoclonal antibody-based assay showed no apparent interference from the presence of human antimouse antibody (HAMA) or the microbiologically inactive vancomycin crystalline degradation product (CDP). The authors conclude that the AxSYM Vancomycin II assay showed satisfactory agreement with other methods tested in this study.


Subject(s)
Anti-Bacterial Agents/blood , Drug Monitoring/standards , Fluorescence Polarization Immunoassay/standards , Vancomycin/blood , Anti-Bacterial Agents/chemistry , Calibration , Chromatography, High Pressure Liquid/standards , Drug Contamination/prevention & control , Drug Monitoring/methods , Enzyme Multiplied Immunoassay Technique/standards , Fluorescence Polarization Immunoassay/methods , Humans , In Vitro Techniques , Infections/blood , Infections/drug therapy , Infections/etiology , Kidney Diseases/blood , Kidney Diseases/complications , Kidney Transplantation/physiology , Linear Models , Reference Values , Reproducibility of Results , Sensitivity and Specificity , Vancomycin/chemistry
9.
Clin Chem ; 42(10): 1645-53, 1996 Oct.
Article in English | MEDLINE | ID: mdl-8855149

ABSTRACT

The major phenytoin metabolite, 5-(p-hydroxyphenyl)-5-phenylhydantoin glucuronide (HPPG), was primarily responsible for the positive bias noted when uremic specimens were assayed with the Abbott TDx Free Phenytoin fluorescence polarization immunoassay. The amount of bias depended on both HPPG and phenytoin concentration, increasing with increases in either concentration. The new Abbott TDx II assays for phenytoin and free phenytoin exhibited no significant cross-reactivity with HPPG and no bias in clinical specimens from uremic patients. Both assays correlated well with Emit-based assays (r >0.98), had CVs of <3.5%, and had minimum detection limits of <0.1 mg/L. Calibration curves were stable for at least 6 weeks. All of the TDx assays cross-reacted with another metabolite, 5-(p-hydroxyphenyl)-5-phenylhydantoin (HPPH), but expected HPPH concentrations are too low to cause a clinically significant bias. The Emit-based phenytoin assay exhibited a significant matrix effect when calibrators were prepared in defibrinated plasma processed to resemble serum.


Subject(s)
Immunoassay/statistics & numerical data , Phenytoin/analogs & derivatives , Phenytoin/blood , Uremia/blood , Calibration , Enzyme Multiplied Immunoassay Technique/standards , Enzyme Multiplied Immunoassay Technique/statistics & numerical data , Fluorescence Polarization , Humans , Immunoassay/standards , Regression Analysis , Sensitivity and Specificity
10.
J Anal Toxicol ; 19(5): 304-6, 1995 Sep.
Article in English | MEDLINE | ID: mdl-7500617

ABSTRACT

Gas chromatography-mass spectrometry (GC-MS) and immunological methods, including the Syva enzyme multiplied immunoassay technique, the Abbott fluorescence polarization immunoassay, and the Roche OnLine immunoassay, were compared for the determination of 11-nor-delta 9-tetrahydrocannabinol-9- carboxylic acid (THCCOOH). The results of all three immunoassays were not in accordance with the GC-MS results in three cases of a 72-specimen panel. Only one false negative was observed using the OnLine immunoassay. The immunological methods compared favorably and are acceptable for detecting the presence of cannabis metabolites in urine. These results support the concept that all immunoassays for cannabinoids should be considered as screening procedures. No concentration correlation between GC-MS and the immunoassays could be established because of the different cross-reactivities of the metabolites.


Subject(s)
Dronabinol/urine , Calibration , Enzyme Multiplied Immunoassay Technique/standards , Fluorescence Polarization Immunoassay/standards , Gas Chromatography-Mass Spectrometry/standards , Humans , Reagent Kits, Diagnostic
11.
Br J Biomed Sci ; 52(2): 102-5, 1995 Jun.
Article in English | MEDLINE | ID: mdl-8520246

ABSTRACT

Enzyme-multiplied immunoassay techniques (EMIT) are now widely used as an effective method for monitoring gentamicin concentrations to ensure adequate serum levels. The current EMIT system was to be superseded by an improved version and both were run in parallel over a nine-month period and the results compared. During this time the systems were evaluated for ease of use, and reproducibility of results. The newer, EMIT produced more consistent calibration results with less kit-to-kit variation, which in turn gave more reproducible results both with patients' sera and the monthly NEQAS quality controls. The newer system was put into routine use at the end of the trial.


Subject(s)
Enzyme Multiplied Immunoassay Technique/standards , Gentamicins/blood , Glucosephosphate Dehydrogenase , Humans
12.
Clin Chem ; 39(8): 1705-12, 1993 Aug.
Article in English | MEDLINE | ID: mdl-8394791

ABSTRACT

The major metabolite of cannabis found in urine, 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid (delta 9-THC), is the compound most often used to calibrate cannabinoid immunoassays. The hydrophobic delta 9-THC molecule is known to adsorb to solid surfaces. This loss of analyte from calibrator solutions can lead to inaccuracy in the analytical system. Because the calibrators remain stable when not used, analyte loss is most probably caused by handling techniques. In an effort to develop an effective means of overcoming adsorption losses, we quantified cannabinoid loss from calibrators during the testing process. In studying handling of these solutions, we found noticeable, significant losses attributable to both the kind of pipette used for transfer and the contact surface-to-volume ratio of calibrator solution in the analyzer cup. Losses were quantified by immunoassay and by radioactive tracer. We suggest handling techniques that can minimize adsorption of delta 9-THC to surfaces. Using the appropriate pipette and maintaining a minimum surface-to-volume ratio in the analyzer cup effectively reduces analyte loss.


Subject(s)
Cannabinoids/urine , Dronabinol/urine , Immunoassay/standards , Adsorption , Calibration/standards , Enzyme Multiplied Immunoassay Technique/instrumentation , Enzyme Multiplied Immunoassay Technique/standards , Enzyme Multiplied Immunoassay Technique/statistics & numerical data , Glass , Humans , Immunoassay/instrumentation , Immunoassay/statistics & numerical data , Plastics , Quality Control , Time Factors
13.
Clin Chem ; 39(7): 1432-4, 1993 Jul.
Article in English | MEDLINE | ID: mdl-8330402

ABSTRACT

We demonstrate here an electrochemical homogeneous enzyme immunoassay for theophylline, which can be performed in hemolyzed, lipemic, and icteric samples. The assay used an unmodified Syva EMIT theophylline kit. One of the enzymatic reaction products, NADH, reacted with 2,6-dichloroindophenol (DCIP) to reduce DCIP to DCIPH2, which was detected electrochemically with flow-injection analysis. The inter- and intraassay coefficients of variation of this manual technique were < 9% at theophylline concentrations of 14 to 34 mg/L. The CVs were 9-15% at low concentrations (6.3 mg/L), which is below the therapeutic range. Analytical recoveries were 91-97% for normal serum and 92-111% for hemolyzed, icteric, or lipemic sera. The measured concentrations (y) were compared with those obtained by the fluorescence polarization immunoassay (x); a scatter plot of the results showed a linear relationship of y = 1.00 x - 0.57 mg/L (r = 0.966, Sy/x = 1.51). This alternative way to measure the serum concentration of theophylline overcomes the shortcomings of spectrophotometric methods, by which it is difficult to measure theophylline in severely hemolyzed, icteric, or lipemic sera.


Subject(s)
Enzyme Multiplied Immunoassay Technique , Theophylline/blood , Bilirubin/blood , Electrochemistry , Enzyme Multiplied Immunoassay Technique/standards , Enzyme Multiplied Immunoassay Technique/statistics & numerical data , Fluorescence Polarization , Hemolysis , Humans , Lipids/blood , Quality Control , Reagent Kits, Diagnostic/standards
14.
Clin Chem ; 39(6): 1089-92, 1993 Jun.
Article in English | MEDLINE | ID: mdl-8504544

ABSTRACT

Immunoassays of cyclosporin A (CsA) have been routinely used to measure CsG. We investigated the cross-reactivities of CsG and its metabolites, as well as the proportion CsG constitutes in relation to total drug measured, for six CsG metabolites (GM1, GM9, GM4N, GM1c, GM1c9, GM19) in the following CsA assays: Sandimmune selective RIA (SS), Sandimmune nonselective RIA (NS), Cyclotrac SP-RIA (CT), fluorescence polarization immunoassay (FPIA), and enzyme immunoassay (EMIT). The cross-reactivity of CsG in these assays was as follows: SS, FPIA, CT, approximately 100%; NS, approximately 40%; EMIT, < 2%. The cross-reactivities of CsG metabolites were investigated in all assays except EMIT and varied among metabolites and assays. The most significant variance was found with the NS assay, where most of the metabolites exhibited cross-reactivities of > 40%. In contrast, in the SS, FPIA, and CT assays, cross-reactivities of < 5% were observed for most of the metabolites. The ranking of cross-reactivities of CsG metabolites in the assays is SS = CT < FPIA < NS. The degree of cross-reactivity did not change significantly when the SS, CT, and FPIA assays were calibrated with CsG instead of CsA--whether parent CsG was present or not. The data suggest that the SS, CT, and FPIA methods would be suitable for the routine monitoring of CsG.


Subject(s)
Cyclosporine/blood , Cyclosporins/blood , Cyclosporins/metabolism , Immunoassay/standards , Cross Reactions , Cyclosporine/immunology , Cyclosporins/immunology , Enzyme Multiplied Immunoassay Technique/standards , False Positive Reactions , Fluorescence Polarization/standards , Humans , Immunoenzyme Techniques/standards , Radioimmunoassay/standards
15.
Br J Biomed Sci ; 50(1): 57-9, 1993 Mar.
Article in English | MEDLINE | ID: mdl-8032295

ABSTRACT

Following suggestions that spurious results occur with EMIT assays--so-called 'fliers'--many laboratories perform such assays in duplicate. In this study, using a Cobas-Bio centrifugal analyser and cost-effective assays developed in the authors' laboratory, complete EMIT assays were run, each with its own calibration curve, on three anti-epileptic drugs using plasma samples, and five drugs of abuse using urine samples. No serious spurious results were obtained. It is suggested that there is no inherent tendency for EMIT assays to produce 'fliers' and that when these occur they may be explained by factors common to many other types of assay.


Subject(s)
Enzyme Multiplied Immunoassay Technique/standards , Autoanalysis , Cost-Benefit Analysis , Enzyme Multiplied Immunoassay Technique/economics , Enzyme Multiplied Immunoassay Technique/instrumentation , Reproducibility of Results
16.
Clin Chem ; 39(1): 104-8, 1993 Jan.
Article in English | MEDLINE | ID: mdl-8419030

ABSTRACT

We evaluated a new EMIT II monoclonal amphetamine/methamphetamine assay for screening human urine by comparing it with the EMIT d.a.u. monoclonal amphetamine/methamphetamine assay and a fluorescence polarization assay. The EMIT II assay has a cutoff of 1 mg/L d-methamphetamine. The EMIT II and EMIT d.a.u. assays were run on a BM/Hitachi 704 analyzer; for the fluorescence polarization assay we used a TDx analyzer. All EMIT II positive samples were also analyzed by the fluorescence polarization assay. We used gas chromatography/mass spectrometry (GC/MS) for confirmation of the presence of amphetamine or methamphetamine. Within-run CVs for the Level 1 (1 mg/L) and Level 2 (3 mg/L) calibrators for the EMIT II assay were 0.47% and 0.53%, respectively. Corresponding between-run CVs were 1.48% and 1.60%, respectively. Of the 1007 samples screened for amphetamines, 50 were positive by the EMIT d.a.u. assay; 21 samples (not a subset of the 50 samples) were positive by the EMIT II assay. However, 19 samples that tested positive by EMIT II also tested positive by the EMIT d.a.u. assay. Subsequent testing of the EMIT II positive samples by the fluorescence polarization assay detected in six positive samples. By means of chiral derivatization wer identified two specimens containing primarily l-isomers of amphetamine and methamphetamine. Sympathomimetic amines were identified in several of the samples not containing amphetamine or methamphetamine.


Subject(s)
Amphetamine/urine , Enzyme Multiplied Immunoassay Technique , Methamphetamine/urine , Reagent Kits, Diagnostic , Antibodies, Monoclonal , Enzyme Multiplied Immunoassay Technique/standards , Enzyme Multiplied Immunoassay Technique/statistics & numerical data , Fluorescence Polarization , Gas Chromatography-Mass Spectrometry , Humans , Quality Control , Reagent Kits, Diagnostic/standards , Reagent Kits, Diagnostic/statistics & numerical data , Substance Abuse Detection
19.
Clin Chem ; 38(11): 2300-6, 1992 Nov.
Article in English | MEDLINE | ID: mdl-1424128

ABSTRACT

In an effort to replace HPLC for whole-blood determination of cyclosporine (CsA), we compared HPLC with radioimmunoassay (RIA; INCSTAR, Cyclo-Trac SP assay), fluorescence polarization immunoassay (FPIA; Abbott TDx), and in-house modified enzyme-multiplied immunoassay technique (EMIT; Syva Co.). For blood samples from 200 various transplant recipients, RIA = 1.262 (HPLC) - 8.16, r = 0.983; FPIA = 1.200 (HPLC) + 19.90, r = 0.981; and EMIT = 1.038 (HPLC) + 11.28, r = 0.985. For segregation by transplant type, RIA, FPIA, and EMIT demonstrated positive biases of 27%, 12%, and 3%, respectively, for liver transplant recipients (n = 50) when compared with HPLC. Heart transplant recipients (n = 50) gave positive bias values of 23%, 14%, and 4% for RIA, FPIA, and EMIT, respectively. Adult renal transplant recipients (n = 50) demonstrated positive bias values of 30%, 31%, and 0% for RIA, FPIA, and EMIT, respectively. For pediatric renal transplant recipients (n = 50), positive biases of 40%, 31%, and 9% were obtained for RIA, FPIA, and EMIT, respectively. We conclude that the modified EMIT represents the best replacement for HPLC.


Subject(s)
Chromatography, High Pressure Liquid , Cyclosporine/blood , Heart Transplantation , Immunoassay , Kidney Transplantation , Liver Transplantation , Chromatography, High Pressure Liquid/statistics & numerical data , Enzyme Multiplied Immunoassay Technique/standards , Fluorescence Polarization Immunoassay/statistics & numerical data , Humans , Immunoassay/statistics & numerical data , Radioimmunoassay/statistics & numerical data , Regression Analysis
SELECTION OF CITATIONS
SEARCH DETAIL
...