Comparison of Procalcitonin Assays on KRYPTOR and LIAISON® XL Analyzers.

Our laboratory performs procalcitonin (PCT) assays on a Brahms KRYPTOR analyzer with the Brahms PCT sensitive Kryptor kit. In this study, we wanted to compare the assays obtained in this way with the ones performed on the LIAISON® XL. From January to May 2017, 171 samples were analyzed, of which 65 from female patients (age: 22-98 years) and 106 from male patients (age: 16-97 years). The PCT determination was performed using the LIAISON® XL and KRYPTOR analyzers, by chemiluminescence (Chemiluminescence immunoassay-CLIA) (LIAISON® BRAHMS PCT® II GEN) and immunofluorescence (Brahms PCT sensitive Kryptor) assay, respectively. For the LIAISON® BRAHMS PCT® II GEN, 52% of the results were placed between 0.0 and 0.5 ng/mL, 18% between 0.5 and 2.0 ng/mL, and 30% between 2.0 and 100 ng/mL; the mean was 4.09 ng/mL, the median 0.456 ng/mL, the maximum value 97.2 ng/mL, and the minimum value 0.02 ng/mL. For the Brahms PCT sensitive Kryptor, 55% of the results were positioned between 0.0 and 0.5 ng/mL, 21% between 0.5 and 2.0 ng/mL, and 24% between 2.0 and 100 ng/mL; the mean was 3.72 ng/mL, the median 0.39 ng/mL, the maximum value 103 ng/mL, and the minimum value 0.01 ng/mL. The mean of the results obtained with the two methods showed no significant differences (3.717 for Kryptor and 4.094 for LIAISON®). PCT assay with Brahms reagents, both on the Kryptor and LIAISON®XL platforms, offers excellent performance in terms of sensitivity and specificity.

Assessment of Access hsTnI 99th percentiles upper reference limits following IFCC recommendations.

BACKGROUND: The detection of an increase and/or decrease of cardiac troponin (cTnI) values, with at least one value above the 99th percentile of the upper reference limit (URL) have a central role in acute myocardial infarction (AMI) diagnosis. The employment of sex specific 99th percentile URLs and High-sensitivity (Hs) assays are recommended. We assessed sex specific 99th percentile URL for Access Hs-cTnI and AccuTnI3+ (Beckman Coulter) using European donor reference population following recent International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) recommendations. METHODS: 300 males and 300 females plasma samples were collected. Both chemiluminescent immunoenzymatic assays were performed on UniCel DxI 800 platform (Beckman Coulter). RESULTS: For Access hsTnI, the observed sex-specific 99th percentile URLs were 5.5 (90% CI: 4.4-7.6) for females and 13.9 ng/L (90% CI: 7.4-17.4) for males. For AccuTnI+3 we could not establish them because the assay couldn't report detectable values of troponin for most of the analyzed samples. CONCLUSION: The sex-specific 99th percentile URLs established for Access hsTnI assay were significantly lower than those declared by the manufacturer caused by the different choice of population selection, age groups and sample types: for those reasons, we maintain the 99th URLs provided by manufacturer.

Improved efficiency and cost reduction in the emergency department by replacing contemporary sensitive with high-sensitivity cardiac troponin immunoassay.

BACKGROUND: Although unquestionable evidence has been provided that high-sensitive (HS) cardiac troponin (cTn) immunoassay outperform the former contemporary-sensitive techniques, some clinicians are still hesitant to implement HS methods in routine clinical practice. This study was hence planned to evaluate the impact of replacing a contemporary-sensitive with HS cTnI immunoassay on hospital and laboratory workload. METHODS: Information on the total number of cTnI tests ordered, total number of blood samples collected, total number of CK-MB tests ordered, number of patients with the first HS-cTnI value below the limit of detection (LoD) and cumulative HS-cTnI values was extracted from the local hospital information system for the semesters before and after the HS method was introduced. RESULTS: Although the total emergency department (ED) visits modestly increased after introducing HS-cTnI, the number of total cTnI tests declined by over 10%. A substantial reduction of single-sample test requests was noted, accompanied by a considerable decline of 3- and 4-sample collections (i.e., -61% and -73%, respectively). A high percentage of patients (27.5%) displayed HS-cTnI values

Analytical imprecision of lactate dehydrogenase in primary serum tubes.

BACKGROUND: Previous studies showed high frequency of duplicate errors of lactate dehydrogenase (LDH) measurement in primary lithium-heparin blood tubes. We hence evaluated imprecision of LDH testing and frequency of replicate errors in primary serum samples centrifuged at two different speeds. METHODS: Serum samples were collected from 15 healthy subjects and centrifuged at either 1300 g or 2000 g. Serum LDH was then measured on Beckman Coulter AU5800 using the International Federation of Clinical Chemistry-recommended procedure. Complete blood cell count of top layer of serum supernatant was also performed using Advia 2120. Imprecision was calculated as coefficient of variation of 10 replicates of each serum tube, whereas the frequency of replicate errors was compared to the desirable specifications for imprecision. RESULTS: Mean LDH values were slightly but significantly higher in serum samples centrifuged at 2000 g rather than 1300 g, whereas blood cell counts were similar. The mean imprecision of LDH was more than double in samples centrifuged at 2000 g than in those centrifuged at 1300 g (8% versus 3%). The percentage of replicate measurements exceeding the desirable specifications was more than threefold higher in samples centrifuged at 2000 g than in those centrifuged at 1300 g (30% versus 9%; P < .001). CONCLUSIONS: The lower frequency of replicated errors in LDH measurement observed using serum samples centrifuged according to manufacturer's instruction suggests that assay precision in serum is better if samples are centrifuged at 1300 g rather than 2000 g when the International Federation of Clinical Chemistry-recommended procedure is used.

Blood collection from intravenous lines: is one drawing site better than others?

OBJECTIVE: Establish whether hemolysis in samples collected from intravenous lines is influenced by catheterization site. METHODS: Blood was collected from all patients (67 total) admitted to the emergency department the same morning, through a 20-gauge catheter placed in a vein of the upper limb directly into an evacuated blood tube. Serum was tested for hemolysis index by multi-wavelength photometric readings. RESULTS: The frequency of hemolyzed specimens was 30% (20/67). Hemolysis rate in median cephalic and basilic veins (17%) was comparable to that of median anterobrachial vein but lower than cephalic vein (29%; P = 0.01), basilic vein (33%; P < 0.01), and metacarpal plexus veins (75%; P < 0.01). Compared with median basilic and cephalic veins, the relative risk of hemolysis was 1.4 from median anterobrachial vein, 1.6 from cephalic vein, 1.9 from basilic vein, and 4.3 from metacarpal plexus veins. CONCLUSION: Drawing blood from catheters placed distally from median veins carries higher hemolysis risk.

The role of neutrophil gelatinase-associated lipocalin (NGAL) in cerebrospinal fluids for screening of acute bacterial meningitis.

BACKGROUND: Acute bacterial meningitis is a rare but extremely severe disease. The aim of this study was to investigate whether neutrophil gelatinase-associated lipocalin (NGAL) is present and measurable in cerebrospinal fluid (CSF) and if its assessment may be useful for identifying patients with bacterial meningitis. METHODS: Eligible specimens were all consecutive CSFs of patients with suspect acute bacterial meningitis that were referred from the Unit of Infectious Diseases for routine chemical and morphological analysis over a three months period. CSF measurements consisted in NGAL, glucose, and total protein concentrations, along with cell count and differential. RESULTS: Eighty eight CSFs were received throughout the study period, 58 (66%) with CSF findings compatible with bacterial meningitis. The values of white blood cells (WBC), polymorphonuclear (PMN) and mononuclear (MONO) leukocytes, red blood cells (RBC), total proteins, and NGAL were significantly increased in positive CSFs, whereas that of glucose did not significantly differ. A significant correlation was found between CSF concentration of NGAL and CSF values of PMN, WBC, RBC and total proteins, but not with that of glucose and MONO. The concentration of NGAL in CSF showed an area under the curve (AUC) of 0.94 for identifying positive CSFs, with specificity and sensitivity of 1.00 and 0.741 at a diagnostic threshold of 13 ng/mL. CONCLUSIONS: NGAL is present in CSF of patients with bacterial meningitis and its measurement may be helpful for identifying positive CSFs.

Reduction of gross hemolysis in catheter-drawn blood using Greiner Holdex tube holder.

INTRODUCTION: Blood collection through intravenous lines frequently causes spurious hemolysis. Due to specific structure, the tube holder Holdex (Greiner Bio-One GmbH, Kremsmuenster, Austria) is supposed to prevent erythrocyte injury in samples collected from catheters, so that we planned a specific study to support this hypothesis. MATERIALS AND METHODS: Blood was collected from emergency department (ED) patients with 20-gauge catheter. In patients with odd order numbers, first and second tubes were collected with conventional holder (BD Vacutainer One Use Holder, Becton Dickinson, Milan, Italy) and the third with Holdex, whereas in even patients first and second tubes were drawn with Holdex and the third using BD Vacutainer One Use Holder. The first tube was discarded, whereas the second and third were centrifuged and serum was tested for potassium, lactate dehydrogenase (LD) and hemolysis index. RESULTS: The final study population consisted in 60 ED patients. Concentrations of potassium (4.25 vs. 4.16 mmol/L; P = 0.031), LD (498 vs. 459 U/L; P = 0.039) and cell-free hemoglobin (0.42 vs. 0.22 g/L; P = 0.042) were higher in samples collected with BD Vacutainer One Use Holder than with Holdex. The mean bias of cell-free hemoglobin was -0.4 g/L in samples collected with Holdex. Although the frequency of samples with cell-free hemoglobin > 0.5 g/L was identical (17/60 vs. 17/60; P = 1.00), the frequency of those with concentrations > 3.0 g/L was higher using BD Vacutainer One Use Holder than Holdex (4/60 vs. 0/60; P = 0.042). CONCLUSIONS: The use of Holdex for drawing blood from intravenous lines may be effective for reducing gross hemolysis.

Systematical assessment of serum indices does not impair efficiency of clinical chemistry testing: a multicenter study.

OBJECTIVES: Despite manufacturers' claim that systematical assessment of serum indices does not impact on testing efficiency, there is widespread perception that this practice may increase the turnaround time (TAT). A multicenter investigation was planned to verify TAT and performance of serum indices on five different clinical chemistry analyzers. DESIGN AND METHODS: Twenty study samples prepared from pooled sera of outpatients, emergency department, intensive care unit and dialyzed patients were divided in aliquots and shipped to 5 different laboratories. According to local instrumentation (Beckman Coulter AU5800, Roche Cobas 6000, Siemens Dimension Vista 1500, Abbott Architect c 16000 and Ortho Vitros 5.1/FS) and reagents, 13 clinical chemistry parameters were assayed on all study samples, with or without contextual assessment of serum indices. RESULTS: The TAT with assessment of serum indices modestly or even negligibly increased, and varied from -0.2 to +5.0% (i.e., from -3 to +85 s). When using the lowest thresholds for sample acceptability, the agreement of hemolysis index (HI) among different instruments was comprised between 0.62 and 1.00 (all p<0.01), but was higher than 0.80 in only 4/10 cases. The agreement of icteric and lipaemic indices could not be estimated due to the low number of samples exceeding acceptability criteria. CONCLUSIONS: The results of this study confirm that systematical measurement of serum indices does not impair instrument efficiency. The comparison of HI also suggests that major harmonization may be advisable for this measure among different manufacturers and instrumentations.

Evaluation of sample hemolysis in blood collected by S-Monovette using vacuum or aspiration mode.

BACKGROUND: In vitro hemolysis can be induced by several biological and technical sources, and may be worsened by forced aspiration of blood in vacuum tubes. This study was aimed to compare the probability of hemolysis by drawing blood with a commercial evacuated blood collection tube, and S-Monovette used either in the "vacuum" or "aspiration" mode. MATERIALS AND METHODS: The study population consisted in 20 healthy volunteers. A sample was drawn into 4.0 mL BD Vacutainer serum tube from a vein of one upper arm. Two other samples were drawn with a second venipuncture from a vein of the opposite arm, into 4.0 mL S-Monovette serum tubes, by both vacuum an aspiration modes. After separation, serum potassium, lactate dehydrogenase (LD) and hemolysis index (HI) were tested on Beckman Coulter DxC. RESULTS: In no case the HI exceed the limit of significant hemolysis. As compared with BD Vacutainer, no significant differences were observed for potassium and LD using S-Monovette with vacuum method. Significant increased values of both parameters were however found in serum collected into BD Vacutainer and S-Monovette by vacuum mode, compared to serum drawn by S-Monovette in aspiration mode. The mean potassium bias was 2.2% versus BD Vacutainer and 2.4% versus S-Monovette in vacuum mode, that of LD was 2.7% versus BD Vacutainer and 2.1% versus S-Monovette in vacuum mode. None of these variations exceeded the allowable total error. CONCLUSIONS: Although no significant macro-hemolysis was observed with any collection system, the less chance of producing micro-hemolysis by S-Monovette in aspiration mode suggest that this device may be used when a difficult venipuncture combined with the vacuum may increase the probability of spurious hemolysis.

Prevention of hemolysis in blood samples collected from intravenous catheters.

OBJECTIVES: Samples drawn through intravenous catheters are frequently hemolyzed. We planned a prospective, randomized study to establish whether hemolysis in samples drawn from intravenous catheters may be reduced using S-Monovette® tubes collected by manual aspiration as compared with standard vacuum tubes. DESIGN AND METHODS: We studied 52 consecutive patients admitted to the ED. Blood was drawn through a 20-gauge intravenous catheter. A 5.0mL, Becton Dickinson Vacutainer® SST II Plus serum tube was collected and discarded. In the odd group of patients (i.e., n. 1, 3, 5, etc.), a second SST II tube was drawn with vacuum ("BD-V"), followed by a 5.5mL S-Monovette® serum tube collected with manual aspiration ("SD-A") and an identical S-Monovette collected by vacuum ("SD-V"). In the pair group of patients (i.e., n. 2, 4, 6, etc.), the sequence was modified to "SD-A", "SD-V" and "BD-V". Serum was separated and tested for lactate dehydrogenase (LDH), potassium and cell-free hemoglobin. RESULTS: The mean concentration of potassium (+2.7% in BD-V and +1.7% in SD-V, respectively), LDH (+15% in BD-V and +7% in SD-V, respectively) and cell-free hemoglobin was significantly increased when samples were collected with vacuum tubes as compared with manual aspiration. No significant differences were observed between SD-V and BD-V. The frequency of hemolyzed samples was higher when blood was collected with the vacuum as compared with SD-A (i.e., 2%), but did not differ between BD-V and SD-V (i.e., 29 versus 31%; p=0.70). CONCLUSION: S-Monovette can be used with vacuum or aspiration collection. This latter approach allows blood drawing with limited shear stress and less likelihood of generating spuriously hemolysis.

Influence of spurious hemolysis on blood gas analysis.

BACKGROUND: Although the prevalence of hemolyzed samples referred for blood gas analysis is as high as 4%, no studies have assessed the bias introduced by spurious erythrocyte breakdown, nor it is known which parameters are mostly influenced and to what extent. This study was hence planned to assess the influence of spurious hemolysis on venous blood gas analysis. METHODS: Venous blood was collected from nine healthy volunteers in sodium heparin tubes and divided in two aliquots of 3 mL. The former aliquot was mechanically hemolyzed by aspiration with 0.5 mL insulin syringe equipped with 30 gauge needle. One milliliter of all aliquots was tested for hemoglobin, pH, oxygen partial pressure (pO2), partial pressure of carbon dioxide (pCO2), bicarbonate (HCO³â»), oxygen tension at 50% hemoglobin saturation (p50), oxygen saturation (sO2), actual base excess (ABE), carboxyhemoglobin (COHb), methemoglobin (metHb), ionized calcium (Ca²âº) and potassium, on ABL800 flex. The remaining 2 mL of blood were centrifuged, plasma separated and tested for hemolysis index. RESULTS: The concentration of cell-free hemoglobin increased from <0.5 g/L to 8.9±1.5 g/L in hemolyzed aliquots. In hemolyzed blood, significant decreases were found for pH (-0.2%), pO2 (-4.9%), sO2 (-4.9%), COHb (-11%) and Ca²âº (-7.0%), whereas significant increases were observed for pCO2 (+4.1%), HCO³â» (+1.4%) and potassium (+152%). Clinically meaningful bias was found for pO2, pCO2, Ca²âº and potassium. CONCLUSIONS: Spurious hemolysis is likely to introduce meaningful biases in blood gas analysis, hence manufacturers of blood gas analyzers should develop instrumentation capable of identifying interfering substances in whole blood. The presence of spurious hemolysis should also be suspected whenever test results do not reflect the clinics.

Assessment of neutrophil gelatinase-associated lipocalin and lactate dehydrogenase in peritoneal fluids for the screening of bacterial peritonitis.

BACKGROUND: Manual microscopy remains the gold standard for enumeration and classification of nucleated cells in peritoneal fluids, especially for diagnosing bacterial peritonitis. However, this approach carries several drawbacks, so that the use of simple and automated tests may be a viable option for initial screening of peritoneal fluids. MATERIALS AND METHODS: Neutrophil gelatinase-associated lipocalin (NGAL), lactate dehydrogenase (LDH), proteins and glucose were assessed in peritoneal fluids from patients with new onset nonmalignant ascites, along with nucleated cell count and differentiation. RESULTS: One hundred and eleven specimens were analyzed, 26 of which (23%) with polymorphonuclear leukocyte (PMN) count≥250/µL, thus compatible with bacterial peritonitis. The median concentration of LDH and NGAL was 3.4 and 3.7-fold higher in samples with ≥250 PMN/µL. The concentration of proteins was also higher in samples with ≥250 PMN/µL, whereas that of glucose was lower. The PMN count significantly correlated with peritoneal fluid values of LDH (r=0.859), NGAL (r=0.774) and proteins (r=0.268), but not with glucose (r=-0.069). The area under ROC curve was 0.88 for LDH, 0.89 for NGAL and 0.94 for their combination (both tests positive), whereas that of proteins and glucose was 0.80 and 0.71, respectively. Sensitivities and specificities were 0.81 and 0.87 for LDH≥227 U/L, 0.96 and 0.75 for NGAL≥120 ng/mL, 0.77 and 0.95 for their combination. The agreement with PMN count was 0.86 for LDH, 0.80 for NGAL, and 0.91 for their combination. CONCLUSIONS: These results suggest that assessment of NGAL in peritoneal fluids, especially in combination with LDH, may be a reliable approach for screening of bacterial peritonitis in patients with new onset nonmalignant ascites.

Influence of mechanical hemolysis of blood on two D-dimer immunoassays.

Although there is broad information about the influence of spurious hemolysis on several laboratory tests, less is known on the bias produced on D-dimer testing. Four different pools were obtained from primary blood tubes, and each of them was divided into four aliquots. The first nonhemolyzed was centrifuged, the plasma was separated and then tested for hemolysis index and D-dimer. The second (hemolyzed aliquot A), third (hemolyzed aliquot B) and fourth (hemolyzed aliquot C) aliquots were mechanically hemolyzed by aspirating whole blood one, two and three times through a fine needle. The plasma was then separated and tested for hemolysis index and D-dimer. D-dimer was quantified by HemosIL AcuStar D-dimer and HemosIL D-dimer HS for ACL TOP. Undetectable hemolysis was present in aliquot nonhemolyzed (<0.5 g/l), whereas the concentration of cell-free hemoglobin significantly increased from hemolyzed aliquot A (5.5-7.0 g/l hemoglobin) to hemolyzed aliquot B (11.5 and 15.0 g/l hemoglobin) and hemolyzed aliquot C (20-22 g/l hemoglobin). The plasma concentration of D-dimer decreased from aliquots nonhemolyzed to hemolyzed aliquot C, achieving clinical significance fromhemolyzed aliquot A and hemolyzed aliquot B when measured with D-dimer HS for ACL TOP and AcuStar D-dimer, respectively. The decrease with AcuStar D-dimer was -5 ± 3% in hemolyzed aliquot A, -7 ± 3% in hemolyzed aliquot B, and -9 ± 3% in hemolyzed aliquot C, whereas the decrease with D-dimer HS for ACL TOP was -5 ± 3% in hemolyzed aliquot A, -8 ± 3% in hemolyzed aliquot B and -9 ± 3% in hemolyzed aliquot C. The similar trend towards decreasing values observed when measuring D-dimer with chemiluminescent and turbidimetric immmunoassays on four heterogeneous plasma pools suggest that the hemolysis interference is more likely to be biological than analytical. The modest bias observed in samples with frank hemolysis (i.e. cell-free hemoglobin of 11.5 g/l) confirms that both methods are robust against this type of interference, so that test results might be released in the majority of mildly hemolyzed samples.

Evaluation of the analytical performances of the novel Beckman Coulter AU5800.

OBJECTIVES: To assess the analytical performance of Beckman Coulter AU5800. DESIGN AND METHODS: Imprecision, linearity and comparison studies were performed on 12 routine clinical chemistry tests. RESULTS: Imprecision was comprised between 0.4 and 3.0%. The linearity was excellent, with regression coefficients always >1.000. The correlation with analogous tests on Unicell DxC ranged from 0.894 and 1.000. The productivity was 4884 tests/h. CONCLUSIONS: We found optimal balance between performance and productivity for the AU5800.

Influence of mechanical trauma of blood and hemolysis on PFA-100 testing.

Although the appropriate quality of samples is essential for platelet function testing, information is lacking on interference from mechanical trauma of blood and hemolysis on PFA-100 analyzer. Citrated blood collected from nine healthy volunteers was divided into three aliquots. The first aliquot ('A') was processed without further manipulation, whereas the second and third were subjected to mechanical trauma by two ('aliquot B') or four passages ('aliquot C') through a very fine needle (30 gauge) to produce hemolysis and cell trauma mimicking poor sample collection. Samples were tested on PFA-100 and Advia 2120, and plasma then separated and tested for lactate dehydrogenase (LDH) and hemolysis index. Negligible hemolysis was present in aliquot A (hemolysis index 0.2 ± 0.1, cell-free hemoglobin 0-0.5 g/l), whereas an increasing amount was present in aliquots B (hemolysis index of 13.1 ± 1.8, cell-free hemoglobin 6.0-6.5 g/l) and C (hemolysis index 24.0 ± 1.1, cell-free hemoglobin 11.5-12.0 g/l). Increases in LDH, and concomitant reductions in platelet and red blood cell counts were observed in aliquots B and C. In hemolyzed aliquots B, four out of nine samples yielded 'flow obstruction' with both PFA-100 agonist cartridges, whereas the closure times were dramatically prolonged in the remaining five samples. In hemolyzed aliquots C, flow obstruction was recorded in six of nine samples for collagen and ADP and all samples for collagen and epinephrine, whereas closure times of collagen and ADP in the remaining three samples were dramatically prolonged. Mechanical trauma of blood causing hemolysis makes PFA-100 testing unreliable. When flow obstructions are observed, the potential presence of hemolysis should be investigated.

Influence of hemolysis on troponin testing: studies on Beckman Coulter UniCel Dxl 800 Accu-TnI and overview of the literature.

BACKGROUND: Hemolyzed specimens are the leading pre-analytical problem in the laboratory practice, and exert a negative impact on test results. We assessed the reliability of Beckman Coulter UniCel Dxl 800 Accu-TnI testing on hemolyzed specimens. METHODS: Twelve non-hemolyzed K2EDTA-anticoagulated samples displaying Accu-TnI values >0.20 µg/L and nine with values <0.04 µg/L were selected and three aliquots were obtained from each. The first ("#A") was processed without further manipulation, whereas the second ("#B") and third ("#C") were hemolyzed by aspirating anticoagulated blood through a thin needle. Plasma was separated and tested for hemolysis index (HI) and Accu-TnI. RESULTS: As compared with aliquot #A (HI: 0), a progressive increase of hemolysis occurred in aliquots #B (HI: 25) and #C (HI: 45). The concentration of Accu-TnI gradually decreased from aliquots #A (0.89 µg/L, 0.20-20.16 µg/L), to aliquots #B (0.81 µg/L, 95% CI 0.17-18.37 µg/L; p=0.041) and #C (0.78 µg/L, 95% CI 0.15-17.48 µg/L; p=0.026). In 0/12 (aliquots #B) and 3/12 cases (aliquots #C) the percent decrease exceeded 20% variation. The values remained unchanged in nine samples with Accu-TnI <0.04 µg/L. CONCLUSIONS: Accu-TnI values decrease in hemolyzed samples, but the bias might not be clinically significant in samples with hemoglobin <14.5 g/L.