Comparison of simultaneous plasma clearance of 99mTc-DTPA and 51Cr-EDTA: can one tracer replace the other?

Both 99mTc-DTPA and 51Cr-EDTA are widely used to determine glomerular filtration rate (GFR), but few direct comparative studies exist. The shortage of 51Cr-EDTA makes a direct comparison highly relevant. The aim of the study was to investigate if there is any clinically relevant difference between plasma clearance of 99mTc-DTPA and 51Cr-EDTA. Patients ≥18 years of age referred for routine GFR measurement by 51Cr-EDTA were prospectively enrolled. The two tracers (10 MBq 99mTc-DTPA (CaNa3-DTPA) and 2.5 MBq 51Cr-EDTA) were intravenously injected at time zero. A standard 4-sample technique was applied with samples collected at 180, 200, 220 and 240 min, if the estimated GFR (eGFR) was ≥30 mL/min. A comparison of single-sample GFR based on the 200 min sample was also conducted. Fifty-six patients were enrolled in the study. All patients had an estimated GFR >30 mL/min/1.73 m2. No patients suffered from ascites or significant oedema. The mean 51Cr-EDTA plasma clearance was 82 mL/min (range 16-226). The plasma clearances determined by the two methods were highly correlated (r = 0.993). The plasma clearance was significantly higher when measured by 99mTc-DTPA than by 51Cr-EDTA (p = 0.01), but the numerical difference was minimal (mean difference 1.4 mL/min; 95% limits of agreement (LOA) -6.6 to 9.4). The difference between the two methods was independent of the level of renal function. Similar results were found for one-sample GFR. No clinically relevant differences were found between the plasma clearance of 99mTc-DTPA and that of 51Cr-EDTA. Therefore, 99mTc-DTPA can replace 51Cr-EDTA when needed.

Single- versus multiple-sample method to measure glomerular filtration rate.

Background: There are many different ways to measure glomerular filtration rate (GFR) using various exogenous filtration markers, each having their own strengths and limitations. However, not only the marker, but also the methodology may vary in many ways, including the use of urinary or plasma clearance, and, in the case of plasma clearance, the number of time points used to calculate the area under the concentration-time curve, ranging from only one (Jacobsson method) to eight (or more) blood samples. Methods: We collected the results obtained from 5106 plasma clearances (iohexol or 51Cr-ethylenediaminetetraacetic acid (EDTA)) using three to four time points, allowing GFR calculation using the slope-intercept method and the Bröchner-Mortensen correction. For each time point, the Jacobsson formula was applied to obtain the single-sample GFR. We used Bland-Altman plots to determine the accuracy of the Jacobsson method at each time point. Results: The single-sample method showed within 10% concordances with the multiple-sample method of 66.4%, 83.6%, 91.4% and 96.0% at the time points 120, 180, 240 and ≥300 min, respectively. Concordance was poorer at lower GFR levels, and this trend is in parallel with increasing age. Results were similar in males and females. Some discordance was found in the obese subjects. Conclusion: Single-sample GFR is highly concordant with a multiple-sample strategy, except in the low GFR range (<30 mL/min).

Strong impact on plasma protein profiles by precentrifugation delay but not by repeated freeze-thaw cycles, as analyzed using multiplex proximity extension assays.

Background A number of factors regarding blood collection, handling and storage may affect sample quality. The purpose of this study was to assess the impact on plasma protein profiles by delayed centrifugation and plasma separation and multiple freeze-thaw cycles. Methods Blood samples drawn from 16 healthy individuals were collected into ethylenediaminetetraacetic acid tubes and kept either at 4 °C or 22 °C for 1-36 h prior to centrifugation. Plasma samples prepared 1 h after venipuncture were also subjected to two to eight cycles of freezing at -80 °C and thawing at 22 °C. Multiplex proximity extension assay, an antibody-based protein assay, was used to investigate the influence on plasma proteins. Results Up to 36 h delay before blood centrifugation resulted in significant increases of 16 and 40 out of 139 detectable proteins in samples kept at 4 °C or 22 °C, respectively. Some increases became noticeable after 8 h delay at 4 °C but already after 1 h at 22 °C. For samples stored at 4 °C, epidermal growth factor (EGF), NF-kappa-B essential modulator, SRC, interleukin 16 and CD6 increased the most, whereas the five most significantly increased proteins after storage at 22 °C were CD40 antigen ligand (CD40-L), EGF, platelet-derived growth factor subunit B, C-X-C motif chemokine ligand 5 and matrix metallopeptidase 1 (MMP1). Only matrix metallopeptidase 7 (MMP7) decreased significantly over time and only after storage at 22 °C. No protein levels were found to be significantly affected by up to eight freeze-thaw cycles. Conclusions Plasma should be prepared from blood after a limited precentrifugation delay at a refrigerated temperature. By contrast, the influence by several freeze-thaw cycles on detectable protein levels in plasma was negligible.

In Haitian women and preschool children, iron absorption from wheat flour-based meals fortified with sodium iron EDTA is higher than that from meals fortified with ferrous fumarate, and is not affected by Helicobacter pylori infection in children.

Fe fortification of wheat flour was proposed in Haiti to combat Fe deficiency, but Fe bioavailability from fortificants has never been investigated in Haitian women or preschool children, two key target groups. We aimed to investigate the bioavailability of ferrous fumarate (FeFum), NaFeEDTA and their combination from fortified wheat flour. We recruited twenty-two healthy mother-child pairs in Port au Prince, Haiti, for an Fe-absorption study. We administered stable Fe isotopes as FeFum or NaFeEDTA individually in low-extraction wheat flour bread rolls consumed by all participants in a randomised, cross-over design. In a final, identical meal, consumed only by the women, FeFum+NaFeEDTA was administered. We measured Fe absorption by using erythrocyte incorporation of stable isotopes 14 d after consumption of each meal, and determined Fe status, inflammatory markers and Helicobacter pylori infection. Fe absorption (geometric mean was 9·24 (95 % CI 6·35, 13·44) and 9·26 (95 % CI 7·00, 12·31) from FeFum and 13·06 (95 % CI 9·23, 19·10) and 12·99 (95 % CI 9·18, 18·39) from NaFeEDTA in mothers and children, respectively (P<0·05 between compounds). Fe absorption from FeFum+NaFeEDTA was 11·09 (95 % CI 7·45, 17·34) and did not differ from the other two meals. H. pylori infection did not influence Fe absorption in children. In conclusion, in Haitian women and children, Fe absorption from NaFeEDTA was 40 % higher than from FeFum, and the combination FeFum+NaFeEDTA did not significantly increase Fe absorption compared with FeFum alone. In the context of Haiti, where the high costs of NaFeEDTA may not be affordable, the use of FeFum at 60 mg Fe/kg flour may be a preferable, cost-effective fortification strategy.

An LC-MS/MS assay for the quantitative determination of 2-pyridyl acetic acid, a major metabolite and key surrogate for betahistine, using low-volume human K2 EDTA plasma.

Betahistine is widely used for the treatment of vertigo. Owing to first-pass metabolism, 2-pyridyl acetic acid (2PAA, major metabolite of betahistine) was considered as surrogate for quantitation. A specific and sensitive LC-MS/MS method was developed and validated for quantitation of 2PAA using turbo-ion spray in a positive ion mode. A solid-phase extraction was employed for the extraction of 2PAA and 2PAA d6 (IS) from human plasma. Chromatographic separation of analytes was achieved using an ACE CN, 5 µm (50 × 4.6 mm) column with a gradient mobile phase comprising acetonitrile-methanol (90:10% v/v) and 0.7% v/v formic acid in 0.5 mm ammonium trifluoroacetate in purified water (100% v/v). The retention times of 1.15 and 1.17 min for 2PAA and internal standard, respectively, were achieved. Quantitation of 2PAA and internal standard was achieved by monitoring multiple reaction monitoring transition pairs (m/z 138.1 to m/z 92.0 and m/z 142.1 to m/z 96.1, respectively). The developed method was validated for various parameters. The calibration curves of 2PAA showed linearity from 5.0 to 1500 ng/mL, with a lower limit of quantitation of 5.0 ng/mL. The bias and precision for inter- and intra-batch assays were <10%. The developed method was used to support clinical sample analysis.

Food Liking Enhances the Plasma Response of 2-Arachidonoylglycerol and of Pancreatic Polypeptide upon Modified Sham Feeding in Humans.

BACKGROUND: Food palatability increases food intake and may lead to overeating. The mechanisms behind this observation are still largely unknown. OBJECTIVES: The aims of this study were the following: 1) to elucidate the plasma responses of endocannabinoids, N-acylethanolamines, and gastrointestinal peptides to a palatable (sweet), unpalatable (bitter), and sensory-acceptable (tasteless control) food, and 2) to verify whether some of these bioactive compounds can serve as plasma biomarkers of food liking in humans. METHODS: Three puddings providing 60 kcal (35% from proteins, 62% from carbohydrates, and 3% from fats) but with different taste were developed. Twenty healthy subjects (11 women and 9 men; mean age 28 y and BMI 22.7 kg/m(2)), selected because they liked the puddings in the order sweet > control > bitter, participated in a randomized crossover study based on a modified sham feeding (MSF) protocol. Blood samples at baseline and every 5 min up to 20 min after the MSF were analyzed for gastrointestinal peptides, endocannabinoids, and N-acylethanolamines. Thirty minutes after the MSF, energy intake at an ad libitum breakfast was measured. RESULTS: After the MSF, no response was observed in 7 of 9 gastrointestinal peptides measured. The plasma ghrelin concentration at 20 min after the sweet and bitter puddings was 25% lower than after the control pudding (P = 0.04), and the pancreatic polypeptide response after the sweet pudding was 23% greater than after the bitter pudding (P = 0.02). The plasma response of 2-arachidonoylglycerol after the sweet pudding was 37% and 15% higher than after the bitter (P < 0.001) and control (P = 0.03) puddings, respectively. Trends for greater responses of anandamide (P = 0.06), linoleoylethanolamide (P = 0.07), palmitoylethanolamide (P = 0.06), and oleoylethanolamide (P = 0.09) were found after the sweet pudding than after the bitter pudding. No differences in subsequent energy intake were recorded. CONCLUSIONS: The data demonstrated that food palatability influenced some plasma endocannabinoid and N-acylethanolamine concentrations during the cephalic phase response and indicated that 2-arachidonoylglycerol and pancreatic polypeptide can be used as biomarkers of food liking in humans.

Relevance of EDTA carryover during blood collection.

BACKGROUND: The order of draw is regarded as a preanalytical issue to prevent carryover of additives during blood collection. Our objective was to prove the theory of ethylenediaminetetraacetic acid (EDTA) carryover for a closed vacuum system and the influence of EDTA on concentrations of selected biomarkers. METHODS: To test the carryover of EDTA, a blood collection with tripotassium EDTA (K3EDTA) and subsequent non-additive tubes was simulated using distilled water as substitute for blood. EDTA concentrations were measured by tandem mass spectrometry. Then we added increasing concentrations of EDTA to heparinized blood and measured routine biomarkers, thereby simulating a carryover of EDTA whole blood and pure EDTA, respectively. Additionally, we tested for EDTA contamination and biomarker alteration in samples collected from 10 healthy volunteers by a syringe with subsequent transfer into sample tubes. RESULTS: No EDTA contamination was detected in samples collected subsequent to a K3EDTA tube when adhering to guidelines of blood sampling. Magnesium, calcium, and potassium levels were altered by artificial K3EDTA whole-blood contamination as well as when adding 1 µL pure K3EDTA. Iron values were altered at EDTA concentrations of 4.4 mmol/L. All other parameters remained unaffected. A slight EDTA carryover was observed in syringe collection and subsequent transfer into EDTA and heparin tubes, however, without any biomarker alteration. CONCLUSIONS: An EDTA carryover during blood collection using a closed vacuum system is highly unlikely. Even if carryover of EDTA whole blood occurs, an absolute volume larger than 10 µL would be necessary to alter test results. However, contamination of samples with preloaded pure K3EDTA solution by severe neglect of current recommendations in blood collection may significantly alter testing results.

Identification and Characterization of EDTA Test Strip Interfering Substances Using a Digital Color Detector.

BACKGROUND: Rapid test strips for ethylenediaminetetraacetic acid (EDTA) can be used to verify correct specimen types for clinical assays which require, or cannot be performed on, plasma collection tubes containing EDTA anticoagulant. As the test strip reaction chemistry is based on a color change induced by chelation of bismuth from a xylenol orange complex, we hypothesized that any agent capable of chelating bismuth might induce false positive test strip reactivity. The present study was therefore designed to evaluate the potential for test strip interference by chelating agents commonly used in the treatment of trace and heavy metal toxicity. METHODS: A digital color detector mounted on a 3D-printed test strip holder was used to quantitatively assess test strip reactivity and evaluate concentration-response relationships of eight commercially available chelating agents. RESULTS: This approach revealed the following rank-order potency: K2EDTA = Na2EDTA > ethylene glycol tetra-acetic acid (EGTA) > dimercaptosuccinic acid (DMSA) > 2,3-dimercapto-1-propanesulfonic acid (DMPS) > penicillamine (PEN). Both deferoxamine (DEF) and alpha lipoic acid (ALA) were non-reactive at 10 mM concentrations. CONCLUSIONS: These experiments demonstrate that multiple substances can induce EDTA rapid test strip reactivity, but only at concentrations higher than might be expected during therapeutic chelation therapy. These agents are therefore unlikely to cause false positive results in routine clinical laboratory specimens.

Validation of a method for the determination of Aderamastat (FP-025) in K2EDTA human plasma by LC-MS/MS.

Aderamastat (FP-025) is a small molecule, selective matrix metalloproteinase (MMP)-12 inhibitor, under development for respiratory conditions which may include chronic inflammatory airway diseases and pulmonary fibrosis. To support evaluation of the pharmacokinetic parameters of Aderamastat in humans, we developed and validated a high-performance liquid chromatography tandem mass spectrometry (LC-MS/MS) analytical method for the quantification of Aderamastat in human plasma. This assay was validated in compliance with the Food and Drug Administration (FDA) Good Laboratory Practice Regulations (GLP) and European Medicines Agency (EMA) guidelines. K2EDTA human plasma samples were spiked with internal standard, processed by liquid-liquid extraction, and analyzed using reversed-phase HPLC with Turbo Ion Spray® MS/MS detection. Separation was done using a chromatographic gradient on 5 µm C6-Phenyl 110 Å, 50*2 mm analytical column at a temperature of 35 °C. The LC-MS/MS bioanalytical method, developed by QPS Taiwan to determine the concentration of Aderamastat in K2EDTA human plasma, was successfully validated with respect to linearity, sensitivity, accuracy, precision, dilution, selectivity, hemolyzed plasma, lipemic plasma, batch size, recovery, matrix effect, and carry-over. These data indicate that the method for determination of Aderamastat concentrations in human K2EDTA plasma can be used in pharmacokinetics studies and subsequent clinical trials with Aderamastat. Authors declare that, this novel data is not published and not under consideration for publication by another journal than this journal. All data will be made available on request.

A new blood collection device minimizes cellular DNA release during sample storage and shipping when compared to a standard device.

BACKGROUND: Cell-free DNA (cfDNA) circulating in blood is currently used for noninvasive diagnostic and prognostic tests. Minimizing background DNA is vital for detection of low abundance cfDNA. We investigated whether a new blood collection device could reduce background levels of genomic DNA (gDNA) in plasma compared to K(3) EDTA tubes, when subjected to conditions that may occur during sample storage and shipping. METHODS: Blood samples were drawn from healthy donors into K(3) EDTA and Cell-Free DNA™ BCT (BCT). To simulate shipping, samples were shaken or left unshaken. In a shipping study, samples were shipped or not shipped. To assess temperature variations, samples were incubated at 6°C, 22°C, and 37°C. In all cases, plasma was harvested by centrifugation and total plasma DNA (pDNA) assayed by quantitative real-time polymerase chain reaction (qPCR). RESULTS: Shaking and shipping blood in K(3) EDTA tubes showed significant increases in pDNA, whereas no change was seen in BCTs. Blood in K(3) EDTA tubes incubated at 6°C, 22°C, and 37°C showed increases in pDNA while pDNA from BCTs remained stable. CONCLUSIONS: BCTs prevent increases in gDNA levels that can occur during sample storage and shipping. This new device permits low abundance DNA target detection and allows accurate cfDNA concentrations.

51Cr-EDTA plasma clearance in children: One, two, or multiple samples?

ABSTRACT: Plasma disappearance curves using multiple blood samples are a recognized reference method for measuring glomerular filtration rate (GFR). However, there is no consensus on the protocol for this type of measurement. A two-compartment model is generally considered acceptable for the mathematical description of the concentration-time decay curve. The impact of the fitting procedure on the reported GFR has not been questioned.We defined 8 different fitting procedures to calculate the area under the curve, and from this area under the curve, the GFR. We applied the 8 fitting methods (all considering a full concentration-time curve) on the multiple sample data (8 samples) of 20 children diagnosed with Duchenne muscular dystrophy. We evaluated the effect (variability) on the reported GFR from the different fitting methods and compared these results with GFR-values calculated from late samples only (samples after 120 minutes) and from one-sample methods.In 6 out of 20 cases, the fitting methods on the full concentration-time curve resulted in very different reported GFR-values, mainly because some methods were not able to fit the data, or methods resulted in GFR-values ranging from 0 to 120 mL/min. The reported GFR-result therefore strongly depends on the fitting method, making the full concentration-time method less robust than expected. Compared with a consensus reference GFR, the late sample models did not show fitting issues and may therefore be considered as more robust. Also the one-sample methods showed acceptable accuracy.The late sample methods (using 3 time-points) provide robust and reliable methods to determine GFR.

Reliability of 5¹Cr-EDTA plasma and urinary clearance as a measure of residual renal function in dialysis patients.

PURPOSE: In dialysis patients, longer survival is associated with a higher residual renal function. Randomized controlled trials are conducted to clarify how residual renal function can be preserved. However, existing methods for measuring residual renal function are uncertain and there is a need for establishing a standard for measurements of glomerular filtration rate (GFR) in dialysis patients. METHODS: 5¹Cr-EDTA clearances in plasma, urine, and dialysate were evaluated in a sample of 12 hemodialysis and 12 peritoneal dialysis patients. The patients' condition was generally stable, and all patients were investigated twice within 4-10 days. RESULTS: Plasma clearances of 5¹Cr-EDTA for all patients ranged between 2.1 and 30.8 mL/min/1.73 m², whereas urinary 5¹Cr-EDTA clearances ranged from 0.7-20.0 mL/min/1.73 m². This difference was statistically significant (p < 0.001). Week-to-week reproducibility expressed as coefficients of variation were below or equal to 10% for plasma clearances and 13% for urinary clearances in hemodialysis patients and 14% in peritoneal dialysis patients. CONCLUSIONS: This study demonstrated a difference between 5¹Cr-EDTA plasma and urinary clearances in dialysis patients. Plasma clearance of 5¹Cr-EDTA had the best reproducibility. For repeated measurements as in clinical prospective trials, we recommend 5¹Cr-EDTA plasma clearance based on blood sampling at 5 + 24 hours with subtraction of 5¹Cr-EDTA dialysate clearance in peritoneal dialysis patients. Further studies are needed to corroborate our findings.

Detection of circulating microfilariae in canine EDTA blood using lens-free technology: preliminary results.

Dirofilaria immitis causes life-threatening heart disease in dogs, thus screening of dog populations is important. Lens-free technology (LFT) is a low-cost imaging technique based on light diffraction that allows computerized recognition of small objects in holographic images. We evaluated an algorithm capable of recognizing microfilariae in canine whole blood using the LFT. We examined 3 groups of 10 EDTA blood specimens, from dogs with microfilaremia (group A), healthy dogs (B), and dogs with hematologic modifications other than microfilaremia (C). The LFT analyzer photographed repeated series of 5 images of all samples. The algorithm declared a sample positive if a microfilaria was detected on ≥1, ≥2, or ≥3 of the 5 images of a series. Microfilariae were detected visually in the images in 9 of 10 cases in group A; no microfilariae were seen in the images from groups B and C. Of the 30 cases, there were 14, 4, and only 3 false-positives with the 1 of 5, 2 of 5, and 3 of 5 image cutoffs, respectively. There were no false-negatives, regardless of cutoff. LFT seems useful for detecting microfilaria and could have application in clinical pathology.

Development and validation of an LC-MS/MS method for determination of hydroxychloroquine, its two metabolites, and azithromycin in EDTA-treated human plasma.

BACKGROUND: Hydroxychloroquine (HCQ) and azithromycin (AZM) are antimalarial drugs recently reported to be active against severe acute respiratory syndrome coronavirus- 2 (SARS-CoV-2), which is causing the global COVID-19 pandemic. In an emergency response to the pandemic, we aimed to develop a quantitation method for HCQ, its metabolites desethylhydroxychloroquine (DHCQ) and bisdesethylchloroquine (BDCQ), and AZM in human plasma. METHODS: Liquid chromatography tandem mass spectrometry was used to develop the method. Samples (20 µL) are extracted by solid-phase extraction and injected onto the LC-MS/MS system equipped with a PFP column (2.0 × 50 mm, 3 µm). ESI+ and MRM are used for detection. Ion pairs m/z 336.1→247.1 for HCQ, 308.1→179.1 for DHCQ, 264.1→179.1 for BDCQ, and 749.6→591.6 for AZM are selected for quantification. The ion pairs m/z 342.1→253.1, 314.1→181.1, 270.1→181.1, and 754.6→596.6 are selected for the corresponding deuterated internal standards (IS) HCQ-d4, DHCQ-d4, BDCQ-d4, and AZM-d5. The less abundant IS ions from 37Cl were used to overcome the interference from the analytes. RESULTS: Under optimized conditions, retention times are 0.78 min for BDCQ, 0.79 min for DHCQ, 0.92 min for HCQ and 1.87 min for AZM. Total run time is 3.5 min per sample. The calibration ranges are 2-1000 ng/mL for HCQ and AZM, 1-500 ng/mL for DHCQ and 0.5-250 ng/mL for BDCQ; samples above the range are validated for up to 10-fold dilution. Recoveries of the method ranged from 88.9-94.4% for HCQ, 88.6-92.9% for DHCQ, 88.7-90.9% for BDCQ, and 98.6%-102% for AZM. The IS normalized matrix effect were within (100±10) % for all 4 analytes. Blood samples are stable for at least 6 hr at room temperature. Plasma samples are stable for at least 66 hr at room temperature, 38 days at -70°C, and 4 freeze-thaw cycles. CONCLUSIONS: An LC-MS/MS method for simultaneous quantitation of HCQ, DHCQ, BDCQ, and AZM in human plasma was developed and validated for clinical studies requiring fast turnaround time and small samples volume.

Multi-centre observational study of spurious hyperkalaemia due to EDTA contamination.

BACKGROUND: A multi-centre observational study investigating the prevalence of spurious hyperkalaemia due to potassium ethylenediaminetetraacetic acid (kEDTA) contamination. METHODS: Serum EDTA was measured in anonymised serum samples with a serum potassium > 6.0 mmol/L collected over a one month period in five different hospital laboratories. Two of the participating laboratories routinely screen all hyperkalaemic samples for EDTA contamination. RESULTS: EDTA contamination was present in 4.1% (range 1.2%-6.7%) of hyperkalaemic samples. In three laboratories, without routine EDTA screening, 50% "EDTA contaminated" were identified by laboratory staff, the remaining 50% samples were undetected and reported as genuine hyperkalaemia. In these laboratories, EDTA was not measurable in 2 samples reported as "EDTA contaminated". CONCLUSIONS: Spurious hyperkalaemia due to kEDTA contamination is relatively common. Education regarding correct blood collection technique offers the best strategy in preventing EDTA sample contamination. Gross kEDTA contamination is easily identified by laboratory staff in samples with marked unexpected hyperkalaemia and hypocalcaemia. Spurious hyperkalaemia due to modest kEDTA contamination may only be confidently detected by measurement of serum EDTA.

The 51-chromium-labeled ethylenediamine tetra-acetic acid intestinal permeability blood test in healthy Beagle dogs: a comparison between serum and plasma levels.

A 51-chromium-labeled ethylenediamine tetra-acetic acid ((51)Cr-EDTA) permeability blood test was validated as a method to assess damage to the small intestine in dogs. The test was performed by calculating various percentages from sera after an orally ingested dose solution. The aim of the current study was to determine whether the use of serum or plasma had any influence on the results of the test. A test solution with 3.7 megabecquerel (100 µCi) of (51)Cr-EDTA was delivered through an orogastric tube to 13 healthy laboratory Beagle dogs. From each dog, 2 concurrent blood samples were withdrawn from cephalic veins using clotting-factor activator tubes for serum and heparinized tubes for plasma. The samples (n  =  26) were taken at 3 and 5 hr after administration of the test solution. Percentages of the orally ingested dose were then calculated in serum and plasma, and their relationship was assessed using correlation analysis. The mean ± standard deviation percentages in serum and plasma after 3 hr were 0.85 ± 0.43% and 0.88 ± 0.49%, respectively, whereas respective percentages in serum and plasma after 5 hr were 0.78 ± 0.52% and 0.81 ± 0.51%. The combined correlation coefficient between the percentages from the sera and plasma samples was excellent (R  =  0.98). It was concluded that the (51)Cr-EDTA permeability test in blood may be performed using serum or plasma of dogs, and the choice between the 2 samples is one of convenience.

Determination of EDTA in dried blood samples by tandem mass spectrometry avoids serious errors in newborn screening.

Newborn screening programs use whole blood dried on filter paper as the standard specimen. Metabolites are reasonably stable and can easily be sent to screening laboratories by regular mail. The recommended sample collection procedure is to spot native blood without anticoagulants onto the filter paper, because anticoagulants can interfere with the different laboratory methods. However, visual examination of the blood spots cannot always detect contamination. In this study, whole blood was drawn by venous puncture from a healthy volunteer, spiked with the corresponding metabolites and EDTA, and spotted onto filter paper. TSH and 17alpha-hydroxyprogesterone were determined by time resolved fluoroimmunoassays with the AutoDelfia system. Total galactose, biotinidase activity, and galactose-1-phosphate uridyltransferase activity were measured photometrically or fluorometrically. Succinyl acetone was estimated indirectly through the inhibition of porphobilinogen synthase activity (PBGS assay). EDTA, amino acids, and acylcarnitines were converted to the corresponding butyl esters, after extraction with methanol, and analysed by LC-MS/MS. EDTA contamination gives falsely elevated 17-OHP values and falsely reduced TSH and PBGS values. The inclusion of an EDTA determination in routine screening revealed that at least 0.06% of newborn screening samples were contaminated with EDTA. In conclusion, non-conformity during the pre-analytical phase is a source of false positive and false negative screening results. Determination of EDTA from NBS blood spots can reliably identify these samples and prevent screening errors.

EDTA sample contamination is common and often undetected, putting patients at unnecessary risk of harm.

BACKGROUND: Potassium ethylenediaminetetraacetic acid (EDTA) is a sample tube anticoagulant used for many laboratory analyses. Gross potassium EDTA contamination of blood samples is easily recognised by marked hyperkalaemia and hypocalcaemia. However, subtle contamination is a relatively common, often unrecognised erroneous cause of spurious hyperkalaemia. Potassium EDTA contamination may also cause hypomagnesaemia and hypozincaemia. There are, however, no data on the prevalence of EDTA contamination as a cause of hypocalcaemia, hypomagnesaemia and hypozincaemia. METHODS: Following a recent service evaluation, we measure EDTA in serum samples from patients with unexplained hyperkalaemia (serum potassium > 6.0 mmol/l). In addition, over a 1-month period EDTA concentrations were measured in hypocalcaemic (serum adjusted calcium < 2.0 mmol/l), hypomagnesaemic (serum magnesium < 0.7 mmol/l) and hypozincaemic (serum zinc < 11 micromol/l) serum samples. RESULTS: Ethylenediaminetetraacetic acid contamination was detected in 31 samples, nine of which were detected by our routine screening programme. The remaining 22 samples represented 14.3% (19/133) of hypocalcaemic samples, 4.8% (5/104) of hypomagnesaemic samples and 1.4% (2/139) of hypozincaemic samples. A total of 25/31 (80.6%) of patients were re-bled, of which 23/25 (92%) results normalised. CONCLUSIONS: Factitious hyperkalaemia, hypocalcaemia and hypomagnesaemia caused by potassium EDTA contamination in our studies are relatively common, and if unrecognised may adversely affect patient care and waste scarce healthcare resources. Correct order of draw of blood samples, improved education and routine laboratory screening of EDTA are necessary to prevent and identify EDTA contamination.

Increased Expression of Meteorin-Like Hormone in Type 2 Diabetes and Obesity and Its Association with Irisin.

Type 2 diabetes (T2D) is a growing pandemic associated with metabolic dysregulation and chronic inflammation. Meteorin-like hormone (METRNL) is an adipomyokine that is linked to T2D. Our objective was to evaluate the changes in METRNL levels in T2D and obesity and assess the association of METRNL levels with irisin. Overall, 228 Arab individuals were enrolled. Plasma levels of METRNL and irisin were assessed using immunoassay. Plasma levels of METRNL and irisin were significantly higher in T2D patients than in non-diabetic patients (p < 0.05). When the population was stratified based on obesity, METRNL and irisin levels were significantly higher in obese than in non-obese individuals (p < 0.05). We found a significant positive correlation between METRNL and irisin (r = 0.233 and p = 0.001). Additionally, METRNL and irisin showed significant correlation with various metabolic biomarkers associated with T2D and Obesity. Our data shows elevated METRNL plasma levels in individuals with T2D, further exacerbated with obesity. Additionally, a strong positive association was observed between METRNL and irisin. Further studies are necessary to examine the role of these proteins in T2D and obesity, against their ethnic background and to understand the mechanistic significance of their possible interplay.