Should we depend on reference intervals from manufacturer package inserts? Comparing TSH and FT4 reference intervals from four manufacturers with results from modern indirect methods and the direct method.

OBJECTIVES: Correct interpretation of thyroid function tests relies on correct reference intervals (RIs) for thyroid-stimulating hormone (TSH) and free thyroxine (FT4). ISO15189 mandates periodic verification of RIs, but laboratories struggle with cost-effective approaches. We investigated whether indirect methods (utilizing historical laboratory data) could replace the direct approach (utilizing healthy reference individuals) and compared results with manufacturer-provided RIs for TSH and FT4. METHODS: We collected historical data (2008-2022) from 13 Dutch laboratories to re-establish RIs by employing indirect methods, TMC (for TSH) and refineR (for FT4). Laboratories used common automated platforms (Roche, Abbott, Beckman or Siemens). Indirect RIs (IRIs) were determined per laboratory per year and clustered per manufacturer (>1.000.000 data points per manufacturer). Direct RIs (DRIs) were established in 125 healthy individuals per platform. RESULTS: TSH IRIs remained robust over the years for all manufacturers. FT4 IRIs proved robust for three manufacturers (Roche, Beckman and Siemens), but the IRI upper reference limit (URL) of Abbott showed a decrease of 2 pmol/L from 2015. Comparison of the IRIs and DRIs for TSH and FT4 showed close agreement using adequate age-stratification. Manufacturer-provided RIs, notably Abbott, Roche and Beckman exhibited inappropriate URLs (overall difference of 0.5-1.0 µIU/mL) for TSH. For FT4, the URLs provided by Roche, Abbott and Siemens were overestimated by 1.5-3.5 pmol/L. CONCLUSIONS: These results underscore the importance of RI verification as manufacturer-provided RIs are often incorrect and RIs may not be robust. Indirect methods offer cost-effective alternatives for laboratory-specific or platform-specific verification of RIs.

Point-of-care high-sensitivity troponin-I analysis in capillary blood for acute coronary syndrome diagnostics.

OBJECTIVES: Patients with acute coronary syndrome (ACS) should be referred promptly to the hospital to reduce mortality and morbidity. Differentiating between low-risk and high-risk patients remains a diagnostic challenge. Point-of-care testing can contribute to earlier disposition decisions for patients excluded from ACS. This study describes the validation of the Atellica® VTLi. Patient-side Immunoassay Analyzer for high-sensitivity troponin point-of-care (POC) analysis. (The Atellica VTLi is not available for sale in the USA. The products/features (mentioned herein) are not commercially available in all countries. Their future availability cannot be guaranteed). METHODS: A total of 152 patients with acute chest pain admitted at the cardiac emergency department (ED) were included in the study. Capillary blood was compared with a whole blood and plasma sample obtained by venipuncture. All samples were analyzed using the Atellica VTLi Patient-side Immunoassay Analyzer; in addition, plasma was analyzed by a central lab immunoassay analyzer. RESULTS: No significant difference was observed between venous whole blood vs. plasma analyzed by the Atellica VTLi Patient-side Immunoassay Analyzer. The difference between capillary blood and venous blood showed a constant bias of 7.1%, for which a correction factor has been implemented. No clinically relevant differences were observed for the capillary POC results compared to plasma analyzed with a standard immunoassay analyzer. CONCLUSIONS: The Atellica VTLi Patient-side Immunoassay Analyzer for high-sensitivity troponin analysis shows equivalent results for all sample types, including capillary blood. No clinically relevant discordances were observed between capillary POC and central laboratory results. With additional studies, this could pave the way towards rapid testing of high-sensitivity troponin in the ambulance or the general practitioner's office without the need for hospitalization of patients with acute chest pain.

Effects of Monomeric and Oligomeric Flavanols on Kidney Function, Inflammation and Oxidative Stress in Runners: A Randomized Double-Blind Pilot Study.

Nonsteroidal anti-inflammatory drugs are frequently used by athletes in order to prevent musculoskeletal pain and improve performance. In combination with strenuous exercise, they can contribute to a reduction of renal blood flow and promote development of kidney damage. We aimed to investigate whether monomeric and oligomeric flavanols (MOF) could reduce the severity of kidney injuries associated with the intake of 400-mg ibuprofen followed by the completion of a half-marathon in recreational athletes. In this double-blind, randomized study, the original MOF blend of extracts from grape seeds (Vitis vinifera L.) and pine bark (Pinus pinaster L.) or placebo were taken for 14 days preceding the ibuprofen/half-marathon. Urine samples were collected before and after the ibuprofen/half-marathon, and biomarkers of kidney injury, inflammation and oxidative stress were assessed. Intake of MOF significantly reduced the incidence of post-race hematuria (p = 0.0004) and lowered concentrations of interleukin (IL)-6 in the urine (p = 0.032). Urinary neutrophil-associated lipocalin, creatine, albumin, IL-8 and malondialdehyde tended to decrease. The supplementation with MOF in recreational runners appears to safely preserve kidney function, reduce inflammation and promote antioxidant defense during strenuous exercise and intake of a single dose of ibuprofen.

Non-steroidal anti-inflammatory drugs increase urinary neutrophil gelatinase-associated lipocalin in recreational runners.

OBJECTIVES: To study the effects of running with/without the use of pain killers on urinary neutrophil gelatinase-associated lipocalin (uNGAL) and other parameters of kidney function in recreational runners. METHODS: Participants of the 10- and 21.1-km Weir Venloop race were enrolled and their urine samples collected before and after the run. Urine dipstick and other conventional tests used to assess kidney function were performed. The presence of ibuprofen, diclofenac, naproxen, and/or paracetamol was assessed by LC-MS/MS. uNGAL was measured with a two-step chemiluminescent immunoassay. RESULTS: NSAIDs/analgesics were detected in urine of 5 (14.4%) 10-km runners and 13 (28.9%) 21.1-km runners. Only half-marathon participants showed significant increases in uNGAL (pre: 11.7 [7.1-34.3] ng/mL; post: 33.4 [17.4-50.4] ng/mL; P = .0038). There was a significant effect of NSAID/analgesic use on uNGAL increase (F2, 76  = 4.210, P = .004). Post hoc tests revealed that uNGAL increased significantly in runners who tested positive for ibuprofen/naproxen compared to runners who did not use any medications (P = .045) or those who tested positive for paracetamol (P = .033). Running distance had a significant influence on the increase in uNGAL (F1, 53  = 4.741, P < .05), specific gravity (F1, 60  = 9.231, P < .01), urinary creatinine (F1, 61  = 10.574, P < .01), albumin (F1, 59  = 4.888, P < .05), and development of hematuria (χ2 (4) = 18.44, P = .001). CONCLUSIONS: Running distance and use of ibuprofen/naproxen were identified as risk factors for uNGAL increase in recreational runners.

Normocalcemia but still elevated parathyroid hormone levels after parathyroidectomy.

Even when laboratory results at first match with clinical assessment, assay interference should still be on a clinician's mind when later results no longer fit with the patient.

Comparison of 7 Published LC-MS/MS Methods for the Simultaneous Measurement of Testosterone, Androstenedione, and Dehydroepiandrosterone in Serum.

BACKGROUND: Recently, LC-MS/MS was stated to be the method of choice to measure sex steroids. Because information on the mutual agreement of LC-MS/MS methods is scarce, we compared 7 published LC-MS/MS methods for the simultaneous measurement of testosterone, androstenedione, and dehydroepiandrosterone (DHEA). METHODS: We used 7 published LC-MS/MS methods to analyze in duplicate 55 random samples from both men and women. We performed Passing-Bablok regression analysis and calculated Pearson correlation coefficients to assess the agreement of the methods investigated with the median concentration measured by all methods, and we calculated the intraassay CV of each method derived from duplicate results and the CVs between the methods. RESULTS: Median concentrations of testosterone were 0.22-1.36 nmol/L for women and 8.27-27.98 nmol/L for men. Androstenedione and DHEA concentrations were 0.05-5.53 and 0.58-18.04 nmol/L, respectively. Intraassay CVs were 2.9%-10%, 1.2%-8.8%, 2.7%-13%, and 4.3%-16% for testosterone in women, testosterone in men, androstenedione, and DHEA. Slopes of the regression lines calculated by Passing-Bablok regression analysis were 0.92-1.08, 0.92-1.08, 0.90-1.13, and 0.91-1.41 for all testosterone values, testosterone in women, androstenedione, and DHEA. Intermethod CVs were 14%, 8%, 30%, and 22% for testosterone in women, testosterone in men, androstenedione, and DHEA. CONCLUSIONS: In general, the LC-MS/MS methods investigated show reasonable agreement. However, some of the assays show differences in standardization, and others show high variation.

Evaluation of a next generation direct whole blood enzymatic assay for hemoglobin A1c on the ARCHITECT c8000 chemistry system.

BACKGROUND: The utility of HbA1c for the diagnosis of type 2 diabetes requires an accurate, precise and robust test measurement system. Currently, immunoassay and HPLC are the most popular methods for HbA1c quantification, noting however the limitations associated with some platforms, such as imprecision or interference from common hemoglobin variants. Abbott Diagnostics has introduced a fully automated direct enzymatic method for the quantification of HbA1c from whole blood on the ARCHITECT chemistry system. METHODS: Here we completed a method evaluation of the ARCHITECT HbA1c enzymatic assay for imprecision, accuracy, method comparison, interference from hemoglobin variants and specimen stability. This was completed at three independent clinical laboratories in North America and Europe. RESULTS: The total imprecision ranged from 0.5% to 2.2% CV with low and high level control materials. Around the diagnostic cut-off of 48 mmol/mol, the total imprecision was 0.6% CV. Mean bias using reference samples from IFCC and CAP ranged from -1.1 to 1.0 mmol/mol. The enzymatic assay also showed excellent agreement with HPLC methods, with slopes of 1.01 and correlation coefficients ranging from 0.984 to 0.996 compared to Menarini Adams HA-8160, Bio-Rad Variant II and Variant II Turbo instruments. Finally, no significant effect was observed for erythrocyte sedimentation or interference from common hemoglobin variants in patient samples containing heterozygous HbS, HbC, HbD, HbE, and up to 10% HbF. CONCLUSIONS: The ARCHITECT enzymatic assay for HbA1c is a robust and fully automated method that meets the performance requirements to support the diagnosis of type 2 diabetes.

A patient with a high concentration of B-type natriuretic peptide (BNP) but normal N-terminal proBNP concentration: A case report.

OBJECTIVES: A 61-year-old female presented with non-typical chest pain. High levels of plasma B-type natriuretic peptide (BNP; result 3188 ng/L, reference range<100 ng/L, method Abbott Architect) were found, although she did not exhibit dyspnoea or other clinical symptoms of heart failure. Echocardiography did not provide an explanation for the elevated BNP concentrations. In follow-up, the chest pain complaints disappeared but BNP remained elevated at the same levels. The serum N-terminal proBNP (NT-proBNP) concentration appeared to be normal. This led us to doubt the accuracy of the BNP values. DESIGN AND METHODS: Possible interference was investigated with BNP and NT-proBNP assays from different manufacturers, various (auto)antibody tests, sample dilutions, addition of mouse serum and polyethylene glycol (PEG) precipitation. RESULTS: BNP and NT-proBNP concentrations were normal when measured using all other (NT-pro)BNP immunoassays. Serial dilutions of sample and addition of mouse serum did not alter the results. Specific (auto)antibody tests were negative. However, PEG precipitation showed the presence of a high molecular weight immunoreactive protein. CONCLUSIONS: We report a false positive BNP result possibly caused by a macro-BNP. This macro-BNP was only immunoreactive in the Abbott Architect BNP immunoassay. Clinicians should be aware of analytical interference when BNP results are constantly elevated in the absence of (non)cardiac causes.

A multicenter evaluation of dysthyroxinemia in a defined patient cohort.

BACKGROUND: In the region Limburg (The Netherlands) almost all of the five participating laboratories use a different immunoassay platform to determine thyroid stimulating hormone (TSH) and free thryoxine (FT4). With the frequent transfer of patients within the region, harmonization of test result interpretation is necessary. In this study, we investigated dysthyroxinemia classification between participating laboratories and developed procedures for improvement. METHODS: Two ring surveys with an interval of 2 years were performed. Four patient groups (n=100) with different dysthyroxinemia classification were based on biochemical results of the Autodelphia analyzer. Samples were tested in five participating laboratories. In each group the percentage of patients classified with dysthyroxinemia was calculated and differences were analyzed by the Fisher's exact test. RESULTS: After the first survey, the percentage of patients with hyperthyroxinemia was more than 20% lower in three laboratories compared to the other two. Bhattacharya analysis revealed that the upper reference limit of FT4 was 20%-30% too high in two laboratories. Adjustments of reference ranges appeared to be effective in the second survey. The third laboratory reported significantly lower percentages of patients with hyperthyroxinemia in the second survey. New FT4 reference ranges were determined for this laboratory, resulting in adequate classification of hyperthyroxinemia. CONCLUSIONS: This study illustrates the potential of a multicenter evaluation of dysthyroxinemia in a biochemical-defined patient cohort. In particular, classification of hyperthyroxinemia differed between laboratories. Adjustments of reference ranges resulted in better agreement of dysthyroxinemia classification. Even using internal and external quality assurance programs, application of multicenter ring surveys is advised to prevent inadequate reference ranges.

Imprecision and limit of quantification of the new architect 25-hydroxyvitamin D assay.

We examined imprecision and limit of quantification (LOQ) of the new Architect 25-hydroxyvitamin D immunoassay in a multicenter setting. Mean intracenter interlot imprecision varied from 22.0 to 5.0%CV in the concentration range of 16 - 92 nmol/L. LOQs were locally determined as 10.8, 11.2, and 13.5 nmol/L. We conclude that the Architect 25(OH)D assay provides a precise and analytically sensitive method for routine measurement of 25(OH)D.

Comparing tolerability and efficacy of generic versus brand alendronate: a randomized clinical study in postmenopausal women with a recent fracture.

INTRODUCTION: An increasing number of generic alendronate formulations have become available. Although expected to have the same tolerability and efficacy, head-to head comparison of generic and brand alendronate was never performed. Therefore, we compared the tolerability and efficacy of generic and brand alendronate. METHODS: In a randomized double-blinded single centre cross-over study in 37 postmenopausal women (mean age 65.4±6.4 years) with osteoporosis were treated with generic and branded alendronate during 24 (2x12) weeks. Tolerance was evaluated by the Gastro intestinal Symptom Rating Scale (GSRS) and self-reported side effects. Efficacy was assessed by serum bone turnover markers, carboxy terminal telopeptide (CTX) and procollagen type I N-terminal propeptide (PINP). No wash out period was allowed (ethical reasons). Because of possible carry over effect only data of the first 12 weeks were analyzed using linear mixed models. RESULTS: There were no significant differences in overall tolerance (GSRS) between treatment groups. However, for subscale abdominal pain, patients using generic had a significantly higher mean GSRS score at week 4 (estimated mean difference (B): 0.40; 95%CI: 0.05 to 0.74, p = 0.024). The level of bone turnover markers significantly decreased over 12 weeks of follow-up for generic and branded alendronate (p < 0.001). Mean level of CTX was significantly lower with branded at week 4 (B: 121.3; 95%CI: 52.0 to 190.5), but not at week 12 (B: 53.6; 95%CI:-3.7 to 110.9). No significant differences were found for PINP at week 4 or 12. CONCLUSIONS: Bone turnover markers were significantly reduced with branded and generic alendronate. With branded, CTX was significantly lower at 4 weeks. Generic caused significantly higher abdominal pain scores in the first 4 weeks of treatment. Therefore, generic alendronate may not have the same tolerability and efficacy as branded alendronate in the first weeks after starting treatment in patients with a recent fracture. TRIAL REGISTRATION: Dutch Trial Register NTR number 1867 http://www.trialregister.nl/trialreg/admin/rctview.asp?TC=1867.

Suboptimal effect of different vitamin D3 supplementations and doses adapted to baseline serum 25(OH)D on achieved 25(OH)D levels in patients with a recent fracture: a prospective observational study.

OBJECTIVE: Guidelines on the need for dose adaptation of vitamin D3 supplementation according to baseline serum 25(OH)D are inconclusive. The effects of increasing doses of vitamin D3 at lower baseline serum 25(OH)D values on the serum 25(OH)D after 4.2 and 11 months were determined in an observational study. DESIGN: A prospective observational study. METHODS: Out of 1481 consecutive women and men with a recent clinical fracture, 707 had a baseline 25(OH)D level <50 nmol/l and were supplemented with increasing doses of vitamin D3 (400, 800, 1700, and ≥3500 IU/day) according to the lower baseline 25(OH)D. Final analysis was restricted to the 221 participants who had full follow-up data available for 11 months. RESULTS: Serum 25(OH)D ≥50 nmol/l was achieved in 57-76% of patients after 4.2 months and in 73-79% after 11 months. These percentages were similar for all doses (P=0.06 and P=0.91 respectively). The mean achieved 25(OH)D was similar for all dose groups (56.1-64.0 nmol/l after 4.2 months and 60.2-76.3 nmol/l after 11 months). With multivariate analysis, the increase in 25(OH)D (17±32.0 after 4.2 months and 24.3±34.0 nmol/l after 11 months) was dependent on the baseline 25(OH)D (P<0.001), not on supplementation dose, season, age, BMI, or gender. CONCLUSIONS: The increase in serum 25(OH)D was significantly larger with higher vitamin D3 supplementation doses. However, this dose-effect response was mainly explained by the baseline 25(OH)D, not the supplementation dose, with a greater magnitude of response at lower baseline 25(OH)D concentrations. In 21-27% of patients, serum 25(OH)D3 levels did not reach 50 nmol/l after 11 months, at any dose. Further studies are needed to identify possible causes of suboptimal response such as non-compliance, undiagnosed malabsorption syndromes, or variability in cholecalciferol content of the vitamin D supplements.

Low vacuum and discard tubes reduce hemolysis in samples drawn from intravenous catheters.

OBJECTIVES: In-vitro hemolysis is a great challenge to emergency departments where blood is drawn from intravenous catheters (IVCs). Although high quality samples can be obtained by straight needle venipuncture, IVCs are preferred for various reasons. The aim of this study was to identify blood collection practices that reduce hemolysis while using IVC. DESIGN AND METHODS: The study was conducted at an emergency department where blood is drawn in ≥ 90% of patients from IVC. Hemolysis, measured spectrophotometrically, was compared between syringe and vacuum tubes. The following practices were tested in combination with vacuum collection; a Luer-slip adapter, a Luer-lock adapter, discard tubes and low vacuum tubes. Each intervention lasted 1 week and retrieved 154 to 297 samples. As reference, hemolysis was also measured in vacuum tubes retrieved from departments where only straight needle venipuncture is performed. RESULTS: Vacuum collection led to more hemolytic samples compared with syringe tubes (24% versus 16% respectively, p=0.008). No difference in hemolysis was observed between the Luer-slip and the Luer-lock adapter. The use of discard (17% hemolytic, p=0.045) and low vacuum tubes (12% hemolytic, p<0.001) substantially decreased hemolysis. None of the interventions reduced the hemolysis rate to the level observed when drawing blood by straight needle venipuncture (3%, p<0.02). CONCLUSIONS: In summary, both discard and low vacuum tubes reduce hemolysis while drawing blood from IVC. Of these practices the use of a low vacuum tube is preferred considering the less volume of blood and the amount of tubes drawn.

Multicenter comparison study of current methods to measure 25-hydroxyvitamin D in serum.

OBJECTIVES: Measurement of serum 25-hydroxyvitamin D [25(OH)D] is generally considered to be a reliable indicator of vitamin D status. The recent increase in diversity of 25(OH)D assays prompted us to evaluate the performance of chromatographic methods (two in-house ID-LC-MS/MS and HPLC (ClinRep, Recipe)), a protein binding method (Cobas-25(OH)D-total, Roche) and immunochemical methods (Liaison and RIA (Diasorin), iSYS (IDS), ADVIA Centaur (Siemens), and Architect i1000 and i2000 (Abbott)). METHODS: Blood was drawn from randomly selected outpatients (N=60) at one site after informed consent. DEQAS and SRM 972 samples were obtained from the scheme organizer and NIST, respectively. Serum aliquots were prepared, frozen and transported to participating centers. Method comparison was performed according to CLSI-EP9 specifications. RESULTS: With these patient samples, and in comparison with ID-LC-MS/MS, Deming regression parameters slope, intercept and R were found to be within the ranges [0.57-1.07], [-1.7 to 6.9 nmol/L] and [0.88-0.98], respectively. 25(OH)D2 in DEQAS and SRM samples was fully recognized by chromatographic methods, but only partially by protein binding and immunochemical methods. Chromatographic methods, and to a lesser extent the protein binding assay, showed cross-reactivity with 3-epi-25(OH)D3. Agreement of 25(OH)D assays to ID-LC-MS/MS in sorting patients into distinct 25(OH)D categories varied between 53% and 88%. CONCLUSIONS: Significant bias exists between ID-LC-MS/MS and many, but not all, other 25(OH)D assays. The variable response among different assays for 25(OH)D metabolites impedes the use of uniform cut-off values for defining vitamin D status. Our results indicate the need towards further standardizing assays for 25(OH)D measurement.

[Diagnostic approach to hypocalcaemia and hypercalcaemia]. / Laboratoriumdiagnostiek bij hypo- en hypercalciëmie.

A 49-year-old woman was examined for osteoporosis and metabolic bone disease after a low-trauma wrist fracture. Laboratory and additional radiological investigations revealed parathyroid hormone (PTH)-mediated hypercalcaemia caused by a parathyroid adenoma. A second patient, a 65-year-old woman with a history of abdominal complaints and tetany, appeared to have hypocalcaemia. Severe vitamin D deficiency and secondary hyperparathyroidism were detected and the patient was finally diagnosed with coeliac disease. Based on these case studies, we highlight the calcium homeostasis and the role of laboratory evaluation of serum calcium, inorganic phosphate, intact PTH, 25-OH vitamin D, magnesium and 24-hour urinary calcium excretion in the diagnostic work-up for hypocalcaemia and hypercalcaemia.