Utility of Cystatin C-based Equation for the Estimation of Glomerular Filtration Rate in a Pediatric Population.

BACKGROUND: The accurate assessment of kidney function is vital for the early detection of kidney damage. The estimated glomerular filtration rate GFR (eGFR) from serum cystatin C (CysC) and creatinine-based equations are commonly used in clinical practice as an alternative to the invasive measured glomerular filtration rate (mGFR), which is the usually accepted overall best index of kidney function in health and disease. Recently the CKiD under 25 (CkiD U25) equations have been shown to perform well in children and young adults with chronic kidney disease (CKD). In this focused report, we evaluated the performance of the CkiD U25 equations alongside 3 non-race-corrected (NRC) eGFR equations commonly used in pediatrics in our cohort. METHODS: mGFR measured following the intravenous injection of tracer Tc-99mDTPA was retrospectively compared with eGFR from these equations in 57 patients (6 months to 22 years) from different races/ethnicities. Ordinary least squares regression analyses were used to assess correlation between the mGFRs and eGFRs. RESULTS: The average mGFR for this cohort was 84.1 mL/min/1.73 m2. The NRC creatinine equations overestimated eGFR across all groups, with the lowest bias for CKiD U25-creatinine (22.59 mL/min/1.73 m2). The best correlations to mGFR, P30, and lowest biases were the CKiD U25-CysC (0.6281, 80.7%, 3.72 mL/min/1.73 m2) and Schwartz CysC (0.6372, 77.2%, -4.68 mL/min/1.73 m2). CONCLUSIONS: Overall, both CKiD U25-CysC and Schwartz CysC provide a good estimation of mGFR with the CKiD U25-CysC having the overall best performance compared to mGFR in our study.

Validation of a whole blood machine learning strategy for distinguishing between bacterial and viral infection in a pediatric hospital setting.

Similar symptoms between viral and bacterial diseases often make diagnosis difficult. This study assessed the clinical performance of the newly cleared whole-blood Bacterial versus Viral Score assay in our pediatric cohort to the previously validated serum assay and emergency department physician diagnosis. This assay shows excellent agreement (R = 0.997) with the serum assay and has great diagnostic accuracy when compared to physician diagnosis.

Validation of a multi-analyte immunoassay for distinguishing bacterial vs. viral infections in a pediatric cohort.

BACKGROUND: Clinical presentation of viral and bacterial infections or co-infections overlaps significantly. Pathogen identification is the gold standard for appropriate treatment. Recently, FDA cleared a multivariate index test called MeMed-BV that distinguishes viral and bacterial infections based on the differential expression of 3 host proteins. Here, we sought to validate MeMed-BV immunoassay on MeMed Key analyzer in our pediatric hospital following guidelines from the Clinical and Laboratory Standards Institute. METHODS: The analytical performance of the MeMed-BV test was evaluated with precision (intra- and inter-assay), method comparison and interference studies. The clinical performance (diagnostic sensitivity and specificity) of the MeMed-BV test was assessed by conducting a retrospective cohort study (n = 60) using plasma samples from pediatric patients with acute febrile illness who visited the emergency department of our hospital. RESULTS: MeMed-BV showed acceptable intra- and inter-assay precision with a range of < 3 score units in both the high-score bacterial as well as the low-score viral controls. Diagnostic accuracy studies revealed a sensitivity of 94% and specificity of 88% for identifying bacterial infections or co-infections. Our MeMed-BV results showed an excellent agreement (R = 0.998) with manufacturer's laboratory data and compared well with ELISA studies. Gross hemolysis and icterus did not affect the assay, but gross lipemia showed a considerable bias in samples with moderate likelihood of viral infection. Importantly, the MeMed-BV test performed better than routinely measured infection-related biomarkers like white blood cell counts, procalcitonin and C-reactive protein in classifying bacterial infections. CONCLUSION: MeMed-BV immunoassay demonstrated acceptable analytical performance and is reliable for distinguishing viral and bacterial infections or co-infections in pediatric patients. Future studies are warranted to examine the clinical utility, especially with respect to reducing the need for blood cultures and time to treatment for the patient.

Performance evaluation of the XT MicroSlide assay pairs on the Vitros XT 7600 compared to VITROS single microslide assays on Vitros 5600.

Objectives: Pediatric hospitals are always challenged by specimen volumes and thus any innovation in this realm is very welcome. With the introduction of Microslide assay pairs, we aimed to evaluate the analytical performance of the Vitros XT MicroSlide assay pairs on the Vitros XT 7600 compared to single MicroSlides. Design: Performance characteristics included within-run precision, analytical measurable range, method comparison, and interference verification. We compared six XT MicroSlide pairs on the Vitros XT 7600 with twelve corresponding single slide assays on the Vitros 5600 system. Results: The XT MicroSlides on Vitros XT 7600 demonstrated excellent precision, equivalent analytical measurable range, and strong method correlation with single slide assays on Vitros 5600 for most of the assays tested. Within-run CVs of the analytes ranged between 0.32% and 2.93% with between-run CV of less than 8.8% and linearity for all analytes was within the manufacturer's specified range. Interference studies showed comparable effects of hemolysis, lipemia, and bilirubin on both instruments. Conclusions: The XT MicroSlides are comparable to the single MicroSlide assays with improved efficiency, turnaround times and lower sample volumes.

Pandemic-Associated Trends in Measurement of HbA1c in Children with Diabetes Mellitus and Validation of Dried Blood Spot as an Alternative Sample Matrix.

OBJECTIVE: Routine monitoring of hemoglobin A1c (HbA1c) is the standard of care in diabetes mellitus (DM), but adhering to regular laboratory appointments may be challenging when access to care is limited, such as during the initial wave of the COVID-19 pandemic in the United States in 2020. MATERIALS: We evaluated trends in patient encounters and laboratory testing for DM in a pediatric healthcare system from March to September 2019 and during the same period in 2020. RESULTS: Evaluation of 17,367 patient encounters illustrated that the pandemic was associated with significantly fewer in-person office visits and point-of-care HbA1c tests for patients with DM in 2020 relative to 2019. A separate analysis of 7,193 HbA1c results measured by point-of-care testing in the general population found a significant increase in the number of measured HbA1c >14 % in May 2020 relative to 2019, but other measured HbA1c values did not differ. As a means to address lapses in the monitoring of HbA1c due to the pandemic, we evaluated the use of the dried blood spot (DBS) matrix for measurement of HbA1c by the Vitros 5600 chemistry analyzer. DBS HbA1c was well correlated to whole blood (r=0.9889) and exhibited intra- and inter-assay precision from 0.5-3.5%. CONCLUSION: DBS measurement of HbA1c presents a viable alternative to routine in-person laboratory monitoring of DM.

Validation of the newly FDA-approved Buhlmann fCal Turbo assay for measurement of fecal calprotectin in a pediatric population.

OBJECTIVES: Inflammatory bowel disease (IBD) is an increasingly prevalent disorder marked by chronic intestinal inflammation. Fecal calprotectin has emerged as a useful biomarker for differential diagnostics and monitoring IBD activity. We validated the newly FDA-approved fCal Turbo fecal calprotectin assay in our pediatric hospital. DESIGN AND METHODS: The performance of the fCal Turbo assay was assessed on the Vitros 5600 analyzer (Ortho Clinical Diagnostics, USA), including limit of quantitation, linearity, precision, and interference studies. Method comparison was performed with 20 fecal samples with the Buhlmann fCal ELISA, and reference range verification was performed with 33 fecal samples. RESULTS: The fCal Turbo assay on the Vitros 5600 was linear between 33.1 and 14,182.5 â€‹µg/g, with dilution studies extending the range to 33.1-22,000 â€‹µg/g, Reproducibility of the assay met acceptability criteria, with intra-assay CV of 0.3-3.2% and inter-assay CV of 5.2-8.9%. Interference studies identified acceptable thresholds for protein, bilirubin, and lipids. We verified a reference range of 33.1-60 â€‹µg/g in our patient population. Deming regression identified acceptable correlation with minor positive bias (2.7%) between the fCal Turbo and fCal ELISA methods. CONCLUSIONS: The fCal Turbo assay performs well on the Vitros 5600 analyzer in our patient population, with the assay being easy to use in our routine chemistry workflow. We anticipate that the fCal Turbo assay will be useful as a rapid screening method for differential diagnostics and disease monitoring of IBD in our patient population.

Clinical Validation and Performance Evaluation of the Automated Vitros Total Anti-SARS-CoV-2 Antibodies Assay for Screening of Serostatus in COVID-19.

OBJECTIVES: Evaluation of serostatus against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has emerged as an important tool in identification of exposure to coronavirus disease 2019 (COVID-19). We report on the validation of the Vitros Anti-SARS-CoV-2 Total (CoV2T) assay for qualitative serologic testing of SARS-CoV-2 antibodies. METHODS: We performed validation studies according to Commission of Office Laboratories Accreditation guidelines, using samples previously tested for SARS-CoV-2 by reverse transcription-polymerase chain reaction (RT-PCR). We evaluated precision, analytical interferences, and cross-reactivity with other viral infections; evaluated concordance with molecular and other serologic testing; and evaluated seroconversion. RESULTS: The Vitros CoV2T assay exhibited acceptable precision and did not exhibit cross-reactivity with other acute respiratory virus infections. The CoV2T assay exhibited 100% negative predictive agreement (56/56) and 71% positive predictive agreement (56/79) with RT-PCR across all patient samples and was concordant with other serologic assays. Concordance with RT-PCR was 97% more than 7 days after symptom onset. The CoV2T assay was robust to icterus and lipemia but had interference from significant hemolysis. CONCLUSIONS: The Vitros CoV2T assay was successfully validated in our laboratory. We anticipate it will be a useful tool in screening for exposure to SARS-CoV-2; however, the use of the CoV2T and other serologic assays in the clinical management of patients with COVID-19 is unknown and must be evaluated in future studies.

Validation of the Procalcitonin Assay on the Abbott Architect i1000.

BACKGROUND: Procalcitonin (PCT) is an emerging biomarker for detecting sepsis. Recently, the US Food and Drug Administration cleared the expanded use of this biomarker for guiding clinicians regarding antibiotic treatment. To our knowledge, there are no published method validations for the Abbott Architect PCT assay. This article will discuss the process of method validation of the B·R·A·H·M·S PCT assay on the Abbott Architect platform. METHODS: We studied the precision, accuracy, and linearity of the Architect method following the guidance of the Clinical and Laboratory Standards Institute EP5-A2 document. Furthermore, we also tested the impact of major sources of interference from hemolysate, lipoproteins, and bilirubin. To validate the Architect method, we compared patients' serum PCT measurements with our previously established Mini VIDAS (bioMerieux) PCT assay. RESULTS: Statistical analysis showed that the 2 assays have good correlation (r > 0.99), slope of 1.023, and intercept of -0.760. The calculated bias is -7.435%. The Architect method showed good precision with %CV < 3.5% for both interassay and intraassay compared with %CV < 6.5% for Mini VIDAS, which was previously determined at our institution. No bias >10% was observed with the Architect method when pooled serum samples were spiked with interferants. The turnaround time for both platforms was the same (20 min); however, in contrast with Mini VIDAS, the Architect system has automated pipetting of samples and can perform multiple assays simultaneously. CONCLUSION: These results showed that the Architect B·R·A·H·M·S PCT assay has analytical characteristics conducive for diagnostic use in clinical laboratories. Our method validation report will be beneficial for other institutions to adapt this assay on existing Abbott Architect i1000 immunoassay analyzers.

Efficacy of two vitamin D immunoassays to detect 25-OH vitamin D2 and D3.

OBJECTIVES: Assessment of Vitamin D status by measurement of 25-Hydroxyvitamin D (25-OH-D) is widely performed by immunoassay. Yet, the ability of these assays to detect Vitamin D2 (as 25-OH-D2) or Vitamin D3 (as 25-OH-D3) varies. It is important to recognize the ability of an assay to quantitate either form of 25-OH-D to evaluate Vitamin D status of supplemented patients. We evaluated detection of 25-OH-D2 and 25-OH-D3 by two assays in our medical center. DESIGN AND METHODS: The Abbott Architect i1000 SR 25-OH Vitamin D assay and Roche Cobas 8000 Vitamin D assay were compared for their recovery of 25-OH-D2 or D3 from spiked serum samples. Samples with known endogenous concentrations of 25-OH-D2 or D3 by LC-MS/MS were also measured to calculate bias between our assays and LC-MS/MS. RESULTS: Recovery of 25-OH-D3 in spiked samples was similar by Architect (84-87%) and Cobas (90%). Recovery of 25-OH-D2 was lower than 25-OH-D3, and was poorer by Architect (37-40%) than by Cobas (69-71%). In measurement of samples with known 25-OH-D concentrations, performance of Architect and Cobas assays was similar for 25-OH-D3. However, at concentrations >50 nmol/L 25-OH-D2, the Architect assay exhibited large average negative bias (-27%). CONCLUSIONS: While the Architect and Cobas assays performed similarly in detection of 25-OH-D3, the Architect assay was significantly poorer at detecting 25-OH-D2 than Cobas, with poorer recovery and significant negative bias at higher concentrations of 25-OH-D2. This agrees with other studies, and indicates that caution should be used in interpreting Architect 25-OH-D results in patients supplemented with Vitamin D2.

Performance of Calculated and Directly Measured Low-Density Lipoprotein Cholesterol in a Pediatric Population.

OBJECTIVES: An assessment of methods for the accurate measurement of low-density lipoprotein cholesterol (LDL-C) at decreased concentrations has not yet been carried out. We evaluated the performance of the Friedewald equation, a direct enzymatic assay, and a novel equation for determining LDL-C levels in a pediatric population with elevated triglycerides and reduced LDL-C levels. METHODS: LDL-C concentrations of 127 pediatric patients were determined by the Friedewald equation, a direct enzymatic assay, and a novel equation. The bias of each approach was assessed at selected LDL-C cutoffs and after stratifying samples by triglyceride content. The concordance of each approach, relative to the reference method, was determined at LDL-C cut-points of less than 70, 70 to 99, and 100 to 129 mg/dL. RESULTS: The Friedewald equation substantially underestimated pediatric LDL-C concentrations below 100 mg/dL in the presence of elevated triglycerides. The Ortho Clinical Diagnostics (Raritan, NJ) direct LDL assay was positively biased at low LDL-C levels. The novel equation most effectively reduced the bias of the Friedewald equation at all LDL-C concentrations and increased the concordance of sample classification to the reference method. CONCLUSIONS: The novel equation should be used for accurate measurement of pediatric LDL-C when the concentration is below 100 mg/dL in the presence of elevated triglycerides (150-399 mg/dL).

Validation of an Automated Assay for Levetiracetam (Keppra) on Vitros 5600.

OBJECTIVE: Levetiracetam (also known as Keppra™) is an antiepileptic drug that has been demonstrated as an effective adjunctive therapy in the treatment of partial onset of seizures, primary generalized tonic-clonic seizures, and myoclonic seizures. The aim of our study was to validate an automated quantitative immunoassay for levetiracetam at Texas Children's Hospital. METHOD: We validated the analytical performance of ARK™ Levetiracetam Assay on an Ortho Clinical Diagnostic Vitros 5600 Analyzer at Texas Children's Hospital. Analytical performance parameters included precision, linearity, sensitivity, accuracy, and effect of common interferents (free hemoglobin, bilirubin, triglycerides). We also tested common drug interferents on the ARK Levetiracetam Assay. RESULTS: The assay showed good precision with <4% coefficient of variation (%CV) for intraassay and <7% for interassay precision, respectively. The assay was linear across the measurement range (0.0-100.00 µg/mL). No significant effect was seen with common interferents or commonly co-used drugs. CONCLUSIONS: The ARK Levetiracetam Assay on Ortho Clinical Diagnostic Vitros 5600 can be used for routine determination of levetiracetam for conducting therapeutic drug monitoring and optimizing individual dosage regimen.

Effect of interference from hemolysis, icterus and lipemia on routine pediatric clinical chemistry assays.

BACKGROUND: Clinical laboratory assays can be affected by interferents like hemoglobin (Hb), lipids and bilirubin. We evaluated the effect of these interferences on pediatric samples for different chemistry assays. Further we established cut-off indices above which these interferences confound sample results. METHODS: Three separate serum pools were spiked with increasing concentrations of hemolysate or intralipid or bilirubin and different analytes were analyzed. The Hemolysis-(H), Lipemia-(T) and Icterus-(I) indices were measured on Vitros 5600. Analytes affected by lipemia were treated with LipoClear ® and re-analyzed. All the measured analytes were compromised by gross hemolysis (H-Index >1000). RESULTS: Except lipase and magnesium (Mg(++)), all other analytes were affected by moderate (H-Index >250) and significant hemolysis (H-Index >500). Low estradiol levels showed a significant effect at severe icterus (I-Index >20.0). C3, C4, Ceruloplasmin, Haptoglobin, Immunoglobulins (Ig) and Vitamin D were significantly affected by moderate (T-Index >100) and severe (T-Index >500) lipemia. LipoClear ® treatment significantly attenuated the lipemic interference on the above analytes except for C3, C4, and IgG. CONCLUSIONS: Accurate reporting of pediatric samples for the analytes affected by common interferences will lead to better clinical interpretation.

Implementation of a quality improvement program to improve sweat test performance in a pediatric hospital.

CONTEXT: All positive screening of newborns for cystic fibrosis using the dried blood spot 2-tiered immunoreactive trypsinogen/DNA method requires subsequent sweat chloride testing for confirmation. Obtaining an adequate volume of sweat to measure chloride is a challenge for many cystic fibrosis centers across the nation. The standard for patients older than 3 months is less than 5% quantity not sufficient (QNS) and for patients 3 months or younger is less than 10% QNS. OBJECTIVE: To set up a quality improvement (QI) program for sweat testing to improve QNS rates using the Wescor Macroduct (Wescor, Inc, Logan, Utah) method at Texas Children's Hospital's laboratory, Houston, Texas. DESIGN: Single-center study. RESULTS: Quantity not sufficient rates were evaluated for 4 months before and 8 months after implementation of the QI program for patients aged 3 months or younger and those older than 3 months. The QI program included changes in technician training, service, site of collection, mode of collection, weekly review, and forms to screen patients for medications that may alter sweat production. A marked improvement was observed in the rates of QNS, which declined considerably from 16.7% to 8.5% (≤3 months old) and from 9.3% to 2.2% (>3 months old) after implementation of the QI initiative in both age categories. CONCLUSION: This report demonstrates the effectiveness of the QI program in significantly improving QNS rates in sweat chloride testing in a pediatric hospital.