Particle-enhanced Turbidimetric Inhibition Immunoassay Research Articles

Three-minute nebulization of gentamicin in healthy dogs results in therapeutic concentrations in bronchoalveolar lavage fluid while remaining below the toxic range values in blood.

To determine and compare the concentration of gentamicin in the lower airways and serum of healthy spontaneously breathing dogs after nebulization with 5% undiluted gentamicin during 3 versus 10 minutes. 10 healthy experimental Beagles. This was a prospective crossover study. A standardized bronchoalveolar lavage (BAL) procedure was performed in each dog after 1 week of administration of each of 2 different gentamicin nebulization protocols separated by a 1-week washout period. The 2 protocols consisted of nebulization of 5% undiluted gentamicin (50 mg/mL) twice daily either during 10 minutes per session (± 95 mg; 10-minute protocol) or 3 minutes per session (± 28 mg; 3-minute protocol). BAL fluid (BALF) was obtained under general anesthesia using a bronchoscope within 15 minutes after administration of the last nebulization. Blood was collected within 5 minutes after BALF collection. BALF and serum gentamicin concentrations were determined by particle-enhanced turbidimetric inhibition immunoassay. Concentrations between protocols were compared using a paired t test. Both BALF and serum gentamicin concentrations were higher after the 10-minute protocol compared with the 3-minute protocol (mean ± SD: 2.41 ± 0.87 mg/L vs 1.25 ± 0.31 mg/L, P = .001; and 1.02 ± 0.59 mg/L vs 0.31 ± 0.24 mg/L, P < .0001 in BALF and serum, respectively), while the BALF-to-serum ratio did not differ between the protocols (3.75 [1.37 to 5.75] (median [IQR]) in the 3-minute protocol vs 2.48 [2.02 to 2.67] in the 10-minute protocol; P = .754). A 3-minute nebulization of gentamicin seems to achieve sufficient concentrations of gentamicin in the BALF to have good efficacy against aminoglycoside-sensitive bacteria while remaining below the toxic range values in blood.

Using Residual Blood from the Arterial Blood Gas Test to Perform Therapeutic Drug Monitoring of Vancomycin: An Example of Good Clinical Practice Moving towards a Sustainable Intensive Care Unit.

Regarding sustainability in the intensive care unit (ICU), there is increasing interest in reducing material waste and avoiding unnecessary procedures. Therapeutic drug monitoring (TDM) of vancomycin, using a dedicated tube, is standard clinical care during treatment with vancomycin. Furthermore, in the ICU, on a daily basis, arterial blood gas (ABG) tests are frequently performed throughout the day. After analysis, a variable volume of blood is discarded. Lithium heparin (LiHep) syringes for ABG tests differ from normally used dipotassium ethylenediaminetetraacetic acid (K2EDTA) tubes. The primary objective was to compare both containers and validate the use of LiHep syringes. Secondary objectives were to evaluate the potential impact on saving materials, nursing time, and costs when implementing vancomycin TDM via LiHep syringes. Vancomycin analysis from sampling in lithium heparin (LiHep) syringes for ABG tests was validated and compared with the concentrations from conventional sampling in K2EDTA tubes. For method comparison, a Bland-Altman plot and Deming regression analysis were performed. The method was validated for inter- and intra-day precision and accuracy. Vancomycin was analyzed by means of the validated method using a particle-enhanced turbidimetric inhibition immunoassay (PETINIA) autoanalyzer. Furthermore, an analysis was conducted to evaluate the potential impact of implementing vancomycin sampling via ABG tests on savings in materials, nursing time, and costs. From 18 patients, 24 plasma samples in both K2EDTA tubes and LiHep syringes were obtained and compared. The mean relative difference between the two containers was -2.0% (-3.0 to -0.93%). Both the Deming regression analysis and the Bland-Altman plot met the acceptance criteria. Potentially, over 1000 blood draws and accompanying materials and packaging can be saved when vancomycin samples are obtained by means of scavenged LiHep syringes. The vancomycin analysis for LiHep syringes showed a total interday precision of 1.95% and an accuracy of 99.7%. The total intraday precision was 2.22%, and the accuracy was 99.2%. Accuracy and precision values were within the acceptance criteria of recovery 85 to 115% and ≤15%, respectively. No significant differences were found in vancomycin concentration between the two analyses, and the LiHep analysis was validated for further implementation in clinical care. Residual blood from ABG test samples can be used for TDM of vancomycin, resulting in a potential reduction of materials used and the number of blood draws. These results will contribute to a more sustainable TDM process with benefits for the patient.

Long-Acting Lipoglycopeptides Can Interfere With Vancomycin Therapeutic Drug Monitoring.

Oritavancin and dalbavancin are long-acting lipoglycopeptides with activity against susceptible gram-positive bacteria, including methicillin-resistant Staphylococcus aureus. Though similar in structure to traditional glycopeptide antibiotics like vancomycin, these antibiotics have terminal half-lives >10 days, and, as a result, there is potential for administration of vancomycin to a patient while oritavancin or dalbavancin are still appreciably present in serum. Given the structural similarities, this creates an opportunity for lab assay interference when performing therapeutic drug monitoring for vancomycin. Following higher-than-expected serum vancomycin concentrations in a patient who received both oritavancin and vancomycin within a short time frame, we evaluated the potential for lipoglycopeptide interference with clinical vancomycin assays. Five platforms covering 3 immunoassay technologies were used to quantify vancomycin concentrations in serum spiked with oritavancin or dalbavancin. Oritavancin generated spurious vancomycin concentrations (20%-84% increase) in both enzyme-multiplied immunoassay technique and a particle-enhanced turbidimetric inhibition immunoassay. However, the improper detection of oritavancin was not consistent across all particle-enhanced turbidimetric inhibition immunoassay platforms. Dalbavancin interference was not detected on any of the platforms tested. The interference from oritavancin may result in falsely elevated vancomycin concentrations and, subsequently, inappropriately adjusted vancomycin doses.

P1798MYCOPHENOLIC ACID TROUGH CONCENTRATION AND DOSE ARE ASSOCIATED WITH HEMATOLOGIC ABNORMALITIES BUT NOT REJECTION IN KIDNEY TRANSPLANT RECIPIENTS

Abstract Background and Aims Little is known regarding the safe fixed dose of mycophenolic acid (MPA) for preventing biopsy-proven acute rejection (BPAR) in kidney transplant recipients (KTRs). We investigated the correlation of MPA trough concentration (MPA C0) and dose with renal transplant outcomes and adverse events. Method This study included 79 consecutive KTRs who received MPA with tacrolimus (TAC) and corticosteroids. The MPA C0 of all the enrolled KTRs was measured, which was determined monthly by using particle-enhanced turbidimetric inhibition immunoassay for 12 months, and clinical data were collected at each time point. The clinical endpoints included BPAR, any cytopenia, and BK or cytomegalovirus infections. Results No differences in MPA C0 and dose were observed between KTRs with or without BPAR or viral infections under statistically comparable TAC concentrations. MPA C0 was significantly higher in patients with leukopenia (P = 0.021) and anemia (P = 0.002) compared with those without cytopenia. The MPA dose was significantly higher in patients with thrombocytopenia (P = 0.002) compared with those without thrombocytopenia. MPA C0 ≥ 3.5 µg/mL was an independent risk factor for leukopenia (adjusted odds ratio [AOR] 3.80, 95% confidence interval [CI] 1.24–11.64, P = 0.019) and anemia (AOR 5.90, 95% CI 1.27–27.51, P = 0.024). An MPA dose greater than the mean value of 1188.8 mg/day was an independent risk factor for thrombocytopenia (AOR 3.83, 95% CI 1.15–12.78, P = 0.029). However, an MPA dose less than the mean value of 1137.3 mg/day did not increase the risk of BPAR. Conclusion Either a higher MPA C0 or dose was associated with an increased risk of cytopenia, but neither a lower MPA C0 nor dose was associated with BPAR within the first year of transplantation. Hence, a reduced MPA dose with TAC and corticosteroids might be safe in terms of reducing hematologic abnormalities without causing rejection.

PETINIA法による血中ミコフェノール酸測定試薬の基準測定法LC-MS/MS法との比較

PETINIA法による血中ミコフェノール酸測定試薬の基準測定法LC-MS/MS法との比較

Mycophenolic Acid Trough Concentration and Dose Are Associated with Hematologic Abnormalities but Not Rejection in Kidney Transplant Recipients.

BackgroundLittle is known regarding the safe fixed dose of mycophenolic acid (MPA) for preventing biopsy-proven acute rejection (BPAR) in kidney transplant recipients (KTRs). We investigated the correlation of MPA trough concentration (MPA C0) and dose with renal transplant outcomes and adverse events.MethodsThis study included 79 consecutive KTRs who received MPA with tacrolimus (TAC) and corticosteroids. The MPA C0 of all the enrolled KTRs was measured, which was determined monthly by using particle-enhanced turbidimetric inhibition immunoassay for 12 months, and clinical data were collected at each time point. The clinical endpoints included BPAR, any cytopenia, and BK or cytomegalovirus infections.ResultsNo differences in MPA C0 and dose were observed between KTRs with or without BPAR or viral infections under statistically comparable TAC concentrations. MPA C0 was significantly higher in patients with leukopenia (P = 0.021) and anemia (P = 0.002) compared with those without cytopenia. The MPA dose was significantly higher in patients with thrombocytopenia (P = 0.002) compared with those without thrombocytopenia. MPA C0 ≥ 3.5 µg/mL was an independent risk factor for leukopenia (adjusted odds ratio [AOR], 3.80; 95% confidence interval [CI], 1.24–11.64; P = 0.019) and anemia (AOR, 5.90; 95% CI, 1.27–27.51; P = 0.024). An MPA dose greater than the mean value of 1,188.8 mg/day was an independent risk factor for thrombocytopenia (AOR, 3.83; 95% CI, 1.15–12.78; P = 0.029). However, an MPA dose less than the mean value of 1,137.3 mg/day did not increase the risk of BPAR.ConclusionEither a higher MPA C0 or dose is associated with an increased risk of cytopenia, but neither a lower MPA C0 nor dose is associated with BPAR within the first year of transplantation. Hence, a reduced MPA dose with TAC and corticosteroids might be safe in terms of reducing hematologic abnormalities without causing rejection.

Comparison of PETINIA and LC-MS/MS for determining plasma mycophenolic acid concentrations in Japanese lung transplant recipients

BackgroundMycophenolic acid (MPA) treatment requires therapeutic drug monitoring to improve the outcome after organ transplantation. The aim of this study was to compare two methods, a particle enhanced turbidimetric inhibition immunoassay (PETINIA) and a reference liquid chromatography tandem mass spectrometry (LC-MS/MS) for determining plasma MPA concentrations from Japanese lung transplant recipients.MethodsPlasma MPA concentrations were determined from 20 Japanese lung transplant recipients using LC-MS/MS and the PETINIA on the Dimension Xpand Plus-HM analyzer.ResultsThe mean MPA concentration measured by PETINIA was significantly higher than that measured by LC-MS/MS (3.26 ± 2.73 μg/mL versus 2.82 ± 2.71 μg/mL, P < 0.0001). The result of the Passing Bablok analysis was a slope of 1.104 (95% confidence interval [CI], 1.036–1.150) and an intercept of 0.229 (95%CI, 0.144–0.315). Bland–Altman analysis revealed PETINIA overestimates plasma MPA concentration by 26.25% and 95%CI from 21.43 to 31.07%.ConclusionThe measurement of MPA by the PETINIA in Japanese lung transplant patients should evaluate the result with attention to positive bias.

Development of a Formula to Correct Particle-Enhanced Turbidimetric Inhibition Immunoassay Values so That it More Precisely Reflects High-Performance Liquid Chromatography Values for Mycophenolic Acid.

BackgroundMycophenolic acid (MPA) concentration measured by homogeneous particle-enhanced turbidimetric inhibition immunoassay (PETINA) may be overestimated due to its cross-reactivity with pharmacologically inactive MPA glucuronide (MPAG), as well as other minor metabolites, accumulated with renal function impairment or co-administered cyclosporine A. In contrast, high-performance liquid chromatography (HPLC) is precise because it can exclude the cross-reactivity. In this study, we assumed HPLC values for MPA (HPLC-MPA) as a reference and aimed to develop a formula correcting PETINA values for MPA (PETINA-MPA) to more precisely reflect HPLC-MPA.MethodsMPA trough concentrations were measured both by HPLC-UV and PETINA in 39 samples issued from 39 solid-organ transplant recipients. MPAG concentrations were also measured using HPLC UV assay. We determined the impacts of renal function and coadministered calcineurin inhibitor on concentrations of MPA and MPAG measured by HPLC. Then, we evaluated the difference between PETINA-MPA and HPLC-MPA. Finally, we develop a formula to reflect HPLC-MPA by using multilinear regression analysis.ResultsMPAG concentration was negatively correlated with estimated glomerular filtration rate (eGFR) (R2 = 0.376, P < 0.001), although MPA was not correlated with eGFR. There were no significant differences in MPA or MPAG concentrations per dose between the patients who were co-administered tacrolimus versus cyclosporine A. Finally, we developed the formulas to reflect HPLC-MPA:Formula 1: Estimated MPA concentration = 0.048 + 0.798 × PETINA‐MPAFormula 2: Estimated MPA concentration = − 0.059 + 0.800 × PETINA‐MPA + 0.002 × eGFRHowever, there was no significant improvement in the coefficient of determination with addition of eGFR in the formula, suggesting that HPLC-MPA can be well predicted by only 1 variable, PETINA-MPA.ConclusionsThis study developed a formula so that PETINA-MPA can be corrected to more precisely reflect HPLC-MPA.

A Spuriously High Phenytoin Level and an Abnormal Lipemic Index: When Aberrant Laboratory Results Lead to a Medical Diagnosis.

An 89-year-old male was admitted for pneumonia and chronic obstructive pulmonary disease (COPD)3 exacerbation. Past medical history was significant for atrial fibrillation, congestive heart failure, COPD, hypertension, diabetes, epilepsy, and mild cognitive impairment. Renal and liver function were within normal limits, with plasma creatinine of 0.86 mg/dL (76 μmol/L) (reference interval 0.68–1.30 mg/dL [60–115 μmol/L]), aspartate aminotransferase of 11 IU/L (reference interval 10–38 U/L), and alanine aminotransferase of 13 IU/L (reference interval 10–80 U/L). Albumin was moderately decreased at 2.8 g/dL (28 g/L) (reference interval 3.4–5.0 g/dL [34–50 g/L]). The patient was on a number of medications, including phenytoin 200 mg in the morning and 160 mg at bedtime. Although he did not exhibit any seizures or other signs of neurological dysfunction during his hospital stay, a phenytoin level was requested to ensure adequate therapy. Total phenytoin was measured on the Siemens Dimension Vista® 1500 platform by particle-enhanced turbidimetric inhibition immunoassay (PETINIA) technology (1) and returned at 28 mg/L (111 μmol/L) (therapeutic range 10–20 mg/L [40–80 μmol/L]), along with error messages of “abnormal assay” and “abnormal reaction.” On-board dilution (4-fold) of the specimen yielded …

Usefulness of mycophenolic acid monitoring with PETINIA for prediction of adverse events in kidney transplant recipients

Background Therapeutic drug monitoring of mycophenolic acid (MPA) is required to optimize the immunosuppressive effect and minimize toxicity. We validated a new particle-enhanced turbidimetric inhibition immunoassay (PETINIA) for the determination of MPA levels and evaluated the relationship of MPA trough level with drug-related adverse events. Methods PETENIA and liquid chromatography-mass spectrometry (LC-MS) were used to determine MPA concentrations from 54 kidney transplant recipients (KTRs). Agreement between PETINIA and LC-MS results was assessed by Passing-Bablok regression and the Bland-Altman plot method. The association of adverse events with MPA trough level obtained by PETINIA was analyzed. Results PETINIA revealed a good agreement with the LC-MS; Regression analysis gave an equation of y = 1.27x − 0.12 (r2 = 0.975, p < 0.001). PETINIA showed a systemic positive bias with a mean difference of 0.66 mg/L compared to LC-MS. However, the magnitude of the positive bias decreased to 0.44 mg/L within the therapeutic range of MPA. Multiple logistic regression showed that MPA trough level determined by PETINIA was an independent risk factor for adverse events (odds ratio 2.28, 95% CI 1.25–4.16, p = 0.007). MPA trough level predicted adverse events with a sensitivity of 77.8% and a specificity of 86.7% using a cut-off level of 5.25 mg/L. Conclusions Good correlation between the two methods indicates that PETINIA is an acceptable method for the monitoring of MPA therapeutic levels. Furthermore, MPA trough level obtained by PETINIA is a useful monitoring tool to minimize toxicity in KTRs.

TO WHAT EXTENT DOES THE VANCOMYCIN ASSAY CONTRIBUTE TO THE VARIABILITY IN VANCOMYCIN CONCENTRATION ESTIMATES IN NEONATES?

*presenting author, supported by ERAWEB II scholarship for postdoctoral program at the KU Leuven, Belgium (2014–2015)IntroductionVancomycin, a glycopeptide antibiotic, is frequently used for late onset sepsis (LOS) and catheter-related infection. Larger inter- and intra-patient variability, combined with a narrow therapeutic index, warrants therapeutic drug monitoring (TDM). However, large inter-individual variability in PK parameters in neonates is documented, only partly explained by covariates such as weight, age or serum creatinine (1,2). In the current study, we focus on the potential impact of between assay differences for vancomycin (3) on the variability in its concentration in a single neonatal intensive care unit (NICU).MethodsVancomycin TDM observations of neonates and young infants treated with intravenous vancomycin, mainly for (suspected) LOS (ie, &gt;72 hours after birth), in the Leuven NICU, Belgium, between June 2011 and December 2014. Our patient population, consists of (pre)term neonates, inborn or transferred, in need of specialized care related to prematurity, infections, perinatal asphyxia, congenital diseases (eg, surgery for cardiopathy, congenital diaphragmatic hernia, or esophageal atresia), or other diseases. Clinical characteristics at birth, as well as characteristics at the moment of TDM were extracted from the patient files. We aimed to document early vancomycin exposure, therefore only first trough levels were included. Serum vancomycin assay was performed either with a particle-enhanced turbidimetric inhibitionimmunoassay method (Siemens Dimension; Dade Behring, Deerfield, Illinois–PETINIA) or with an enzyme multiplied immunoassay technique (Cobas c702; Roche Diagnostics, Basel, Germany–COBAS). The data were analyzed by Chi-square test, t test and Mann-Whitney U test. Linear Mix Model was used to assess significant differences between groups, when adjusting for confounding factors. Data were analyzed in SPSS 20.0 (IBM corp.), p-value &lt;0.05 was significant.ResultsIn total, 564 vancomycin TDM observations, 311 assayed with PETINIA and 253 with COBAS, were included. Both cohorts had comparable clinical characteristics (median [min-max] current weight 2150 [420–5000] grams for PETINIA vs. 2120 [500–5840] grams for COBAS, and median postmenstrual age 35 [25–58] weeks for PETINIA vs. 35 [25–51] weeks for COBAS). We determined the significant difference between the vancomycin concentrations using two different immunoassays: PETINIA vs. COBAS (F=17.971; p&lt;0.001). When adjusting for current body weight and postmenstrual age, the major covariates associated with vancomycin serum trough levels in neonates, the difference in vancomycin concentration between cohorts was statistically significant (F=17.076, p&lt;0.001, F=18.951, p&lt;0.001, respectively). Overall, immunoassays PETINIA and COBAS significantly differed by vancomycin concentrations when adjusting for covariates, and the mean difference for vancomycin concentration was 2.167 mg/l.ConclusionThe present study confirms the impact of assays on the variability in vancomycin concentration in neonates in a single NICU. Comparison between these two immunoassays showed a mean proportional differences &gt;20%. Therefore, it is important to know how the vancomycin is measured when interpreting results, and particularly the transferability of vancomycin results between the laboratories has to be interpreted with caution.

TDMの落とし穴

Therapeutic drug monitoring (TDM) is an essential tool for the optimal use of drugs. Although rare, the drug concentration in blood determined by TDM can be higher or lower than that predicted from the dosage amount and patient information. In such cases, it is necessary to consider the occurrence of false positives and false negatives in the measurement process. Dose adjustment based on the measured blood drug concentration poses potential risks of underdosing and overdosing in patients suspected of false positive and false negative data, respectively. Therefore, pharmacists involved in TDM are required to become familiar with false positives and false negatives in TDM. In particular, the possibility of false positives in TDM of immunosuppressants by the affinity column-mediated immunoassay (ACMIA) method should be considered, while TDM of vancomycin or phenytoin requires attention to false negatives during measurement by the particle-enhanced turbidimetric inhibition immunoassay (PETINIA) method. Moreover, interference from IgM in the measurement process may be of concern in false-negative results; therefore, the measured blood drug concentration needs to be verified when the blood IgM level is high. This mini-review outlines false positives and false negatives that should be considered in TDM and presents a discussion of the relevant literature.

Possible False-Negative Results on Therapeutic Drug Monitoring of Phenytoin Using a Particle Enhanced Turbidimetric Inhibition Immunoassay in a Patient With a High Level of IgM

: In this report, the authors described the unusual case of a patient in whom the plasma phenytoin concentration was unexpectedly not detected on a particle-enhanced turbidimetric inhibition immunoassay (PETINIA) technique, a typical immunoassay for phenytoin. The plasma concentration was measured using PETINIA and high-performance liquid chromatography in a 69-year-old male patient treated with fosphenytoin intravenously at the standard dose for 7 days. Although the plasma concentration of phenytoin was below the limit of detection (<0.5 mcg/mL) on PETINIA after the administration of fosphenytoin, the trough plasma concentration was estimated to be between 5 and 10 mg/L on high-performance liquid chromatography. When the plasma concentrations of IgM and IgG were measured using an enzyme-linked immunosorbent assay, the plasma IgG level was within the reference range, whereas the plasma IgM level was 2-3 times higher than the upper limit of the reference range. We concluded that the PETINIA method yielded a possible false-negative result regarding the phenytoin level in this patient, perhaps because of some hindrance to the measurement process by IgM. This case suggests that false-negative results should be considered when therapeutic drug monitoring reveals abnormally low values using PETINIA and that it is necessary to evaluate the plasma IgM level.

Digoxin Immunoassays on the ARCHITECT i2000SR and ARCHITECT c8000 Analyzers Are Free From Interferences of Asian, Siberian, and American Ginseng

Asian, Siberian, and American ginseng are known to interfere with serum digoxin measurements using fluorescence polarization technology, Digoxin II and Digoxin III assays (Abbott Laboratories, Green oaks, IL) as well as other digoxin assays. Abbott Laboratories more recently launched two new digoxin assays: iDigoxin, a chemiluminescent microparticle immunoassay for application on the ARCHITECT i1000SR and i2000SR immunoassay analyzers, and cDigoxin, a particle-enhanced turbidimetric inhibition immunoassay for application on the ARCHITECT c4000, c8000, and c1600 clinical chemistry analyzers; and we studied potential interferences of ginsengs with these two assays in vitro. When aliquots of drug-free serum pool treated with activated charcoal were supplemented with extracts of various ginsengs, no significant apparent digoxin values were observed. In addition, when aliquots of the digoxin pool prepared from patients taking digoxin were further supplemented with these ginseng extracts and the digoxin values were re-measured, we observed no statistically significant difference in observed digoxin values compared to the original digoxin value of the pool. These results further establish that relatively new digoxin assays for application on the ARCHITECT analyzers that employ specific monoclonal antibodies against digoxin are free from interferences from Asian, Siberian, and American ginseng.

Mathematical Equations to Calculate True Mycophenolic Acid Concentration in Human Plasma by Using Two Immunoassays With Different Cross‐Reactivities With Acyl Glucuronide Metabolite: Comparison of Calculated Values With Values Obtained by Using an HPLC‐UV Method

Both immunoassays and chromatographic methods are available for therapeutic drug monitoring of mycophenolic acid (MPA). Although chromatographic methods are more precise, immunoassays are widely used in clinical laboratories due to ease of adopting such assays on automated analyzers. We studied the possibility of using mathematical equations to calculate true MPA concentration by accounting for acyl glucuronide cross-reactivities with immunoassays by using two immunoassays with widely different cross-reactivities with the metabolite. We determined MPA concentrations in 20 specimens obtained from transplant recipients using cloned enzyme donor immunoassay (CEDIA) assay and a new particle enhanced turbidimetric inhibition immunoassay (PETINIA) assay. Then we developed mathematical equations to calculate true MPA concentration using values obtained by both immunoassays and reported cross-reactivity of acyl glucuronide with respective immunoassays. Calculated concentrations were compared with values obtained by using a high-performance liquid chromatography combined with ultraviolet detection (HPLC-UV) method. We obtained good correlation between calculated MPA concentrations and corresponding MPA level obtained by using HPLC-UV method. Using x-axis as the MPA concentrations determined by the HPLC-UV method and y-axis as the calculated MPA level, we observed the following regression equation: y = 1.083x - 0.0995 (r = 0.99, n = 20). Mathematical equations can be used to calculate true MPA concentrations using two immunoassays with different cross-reactivities with acyl glucuronide metabolite.

PETINIA Total Phenytoin Assay on the Vista 1500 Analyzer Is Not Suitable for Monitoring Free Phenytoin Concentration if Ultrafiltration Is Performed at Room Temperature

Phenytoin is a strongly protein-bound anticonvulsant requiring free drug monitoring for certain patients. Recently, free phenytoin assay on the TDx analyzer was discontinued by the Abbott Laboratories, but Roche Diagnostics has an assay for the determination of free phenytoin. Because a clinical laboratory may not have an instrument for using Roche free phenytoin assay, we studied the possibility of determining free phenytoin using particle-enhanced turbidimetric inhibition immunoassay (PETINIA) total phenytoin assay on the Vista 1500 analyzer by comparing free phenytoin values obtained by Roche assay and Hitachi 917 analyzer with values obtained by using PETINIA assay. When protein-free ultrafiltrates were prepared at room temperature from 41 different patients taking phenytoin, using Roche assay (x axis) and PETINIA (y axis) assay, the following regression equation was observed: y = 0.8526x + 0.0661 (r = 0.85, n = 41). The regression equation indicated significant negative bias with the reference method. However, when additional 30 specimens were studied where ultrafiltration was performed at 37°C, lesser negative bias was observed as revealed by the regression equation: y = 0.922x - 0.117 (r = 0.90, n = 30). We conclude that PETINIA assay on the Vista 1500 analyzer is not suitable for monitoring free phenytoin concentration if ultrafiltration is performed at room temperature but may be used with caution if ultrafiltration is performed at physiological temperature of 37°C, which is the desirable temperature for preparation of ultrafiltrate.

Two Suspected Cases of Immunoglobulin-Mediated Interference Causing Falsely Low Vancomycin Concentrations with the Beckman PETINIA Method

To describe 2 recent cases of suspected immunoglobulin-mediated interference with the Beckman Coulter particle-enhanced turbidimetric inhibition immunoassay (PETINIA) used to measure vancomycin serum or plasma concentrations and to review the existing literature. A 64-year-old woman with a history of multiple immune-related comorbidities received vancomycin for treatment of a prosthetic joint infection growing coagulase-negative Staphylococcus spp. A 33-year-old man with a history of Felty syndrome received vancomycin for the treatment of methicillin-resistant Staphylococcus aureus pneumonia. Both patients had multiple vancomycin trough concentrations determined using the Beckman Coulter PETINIA method and had measured concentrations reported as less than 4 mg/L despite appropriate vancomycin dosing for their size, age, and organ function. The patients' serum was then tested by alternative methods, which reported vancomycin concentrations consistent with those expected with the patients' dosing regimens. Immunoglobulins are well known for interfering with chemistry assays. It is suspected that high levels of immunoglobulins in these 2 patients interfered with the accurate measurement of serum vancomycin concentrations. An objective causality assessment supported the interactions as being definite and probable, respectively. Two other cases of suspected immunoglobulin interference with vancomycin measurement have been reported in the literature when vancomycin concentrations were initially measured using the Beckman Coulter PETINIA method. In vitro studies support the hypothesis that immunoglobulin interference with vancomycin measurement by this method occurs and is a graded phenomenon. These cases suggest a high likelihood of immunoglobulin-mediated interference with the Beckman Coulter PETINIA method, which results in the reporting of falsely low vancomycin serum concentrations. When vancomycin concentrations do not correlate with what would be expected clinically, clinicians should have a high index of suspicion for potential assay interference.

Comparison of mycophenolic acid concentrations determined by a new PETINIA assay on the Dimension EXL analyzer and a HPLC-UV method

ObjectiveMycophenolic acid requires routine therapeutic drug monitoring. We evaluated the suitability of a new PETINIA (particle enhanced turbidimetric inhibition immunoassay) assay on the Dimension EXL analyzer for monitoring of mycophenolic acid by comparing values obtained by this assay with a HPLC-UV method. Design and methodsMycophenolic acid concentrations determined in sera of 60 organ transplant recipients using high performance liquid chromatography combined with ultraviolet detection (HPLC-UV, reference method) and the new immunoassay on the Dimension RxL analyzer. ResultsThe within and between run precision of the new PETINIA assay was <10%. The assay was linear for a mycophenolic acid concentration up to 30μg/mL. When mycophenolic acid concentrations in 60 transplant recipients obtained by the HPLC-UV (x-axis) method were compared with corresponding values obtained by the PETINIA assay (y-axis), the following regression equation was obtained: y=1.1204 x+0.0881 (r=0.983, n=60). ConclusionsIf PETINIA assay is used for therapeutic drug monitoring of mycophenolic acid, caution must be exercised in interpreting serum mycophenolic acid level due to observed positive bias.

Analytical Validation of a Homogeneous Immunoassay for Determination of Mycophenolic Acid in Human Plasma

Mycophenolic acid (MPA) is an immunosuppression agent for the prophylaxis of organ rejection in patients receiving allogeneic transplants. The drug is administered based in 2 formulations, mycophenolate mofetil (MMF) and enteric-coated mycophenolate sodium (EC-MPS). MPA acts by specific, reversible, uncompetitive inhibition of inosine monophosphate dehydrogenase (IMPDH) and thus blocks the proliferation of both T- and B-activated lymphocytes. Therapeutic drug monitoring (TDM) constitutes an important part of immunosuppressive treatment because of the demonstrated significant intraindividual and interindividual variability of its pharmacokinetic behavior. TDM is required to optimize immunosuppressive efficacy. We present the analytical validation of a homogeneous particle-enhanced turbidimetric inhibition immunoassay (PETINIA) technique for determination of MPA in human plasma, and compare with a homogeneous enzyme immunoassay technique (EMIT; reference method), both methods adapted on a Dimension analyzer (Siemens). We examined 50 human plasma samples from kidney transplant recipients treated with MMF or EC-MPA, which were analyzed simultaneously by both methods. The interassay precision was 5.95% at a concentration of 1.0 μg/mL, 3.47% at 7.5 μg/mL, and 3.75% at 12.0 μg/mL. The bias of PETINIA-MPA for each of the 3 quality control sample was <3.0%. Least squares linear regression yielded an r-value of 0.994 with the following linear regression equation: PETINIA = 0.939 * EMIT − 0.063. Bland-Altman comparison presented a mean negative difference of −0.312 μg/mL (standard deviation [SD], 0.441), namely, −7.6% for PETINIA-MPA. The PETINIA assay for monitoring MPA concentrations is an acceptable method for routine clinical use, with interassay imprecision (% coefficient of variation) ranging from 5.9% to 3.7% below and above the therapeutic concentration range, respectively. In conclusion, MPA-EMIT and PETINIA-MPA methods on Dimension analyzer have a good correlation (r = 0.994), but PETINIA-MPA method demonstrates a negative average difference of −7.6% in comparison with EMIT-MPA method.

Release of Vancomycin and Gentamicin from a Contact Lens Versus a Fibrin Coating Applied to a Contact Lens

To develop a contact lens capable of releasing antibiotics for a minimum of 8 hours for the treatment of bacterial keratitis. Fibrin gel was loaded with vancomycin or gentamicin and then shaped into a curved disc. The disc was then used to coat the surface of a commercial contact lens or was sealed between two lenses. Separate contact lenses were soaked in solutions of vancomycin or gentamicin. The in vitro release kinetics for each system was determined using PBS at 37°C and a particle-enhanced turbidimetric inhibition immunoassay. The bioactivity of the antibiotics released from the fibrin was confirmed by using a microbiological assay. Vancomycin and gentamicin were released at similar rates from soaked contact lenses and a coating of fibrin gel; however, the amounts of antibiotic delivered by the two systems differed considerably. The fibrin coating released over three times more gentamicin but less than one-fifth that of the lenses soaked in vancomycin. When fibrin was encapsulated between two contact lenses, significantly more controlled release was observed. For all systems, bactericidal amounts of vancomycin and gentamicin were released throughout the three-day testing period. As a delivery system, fibrin gel loaded with gentamicin performs better than contact lenses soaked in gentamicin. The opposite is true for vancomycin, where soaked lenses outperform fibrin gel. These systems could potentially be used as a treatment for bacterial keratitis.