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1.  Physical activity - an important preanalytical variable 
Biochemia Medica  2014;24(1):68-79.
The concentration of several biochemical and hematological biomarkers is strongly influenced by a number of preanalytical variables. Several lines of evidence attest that short, middle, and long-term exercise, as well as the relative intensity of physical effort (from mild to strenuous), may influence a broad array of laboratory variables. The amount of extracellular release and clearance from blood of most of these biomarkers is markedly influenced by the biological characteristics of the molecule(s), level of training, type, intensity and duration of exercise, and time of recovery after training. It is hence noteworthy that test results that fall outside the conventional reference ranges in athletes not only may reflect the presence of a given disease, but may frequently mirror an adaptation to regular training or changes that have occurred during and/or following strenuous exercise, and which should be clearly acknowledged to prevent misinterpretation of laboratory data. The aim of this narrative review is to provide an update about the most significant changes of some biochemical and hematological biomarkers in response to physical exercise, for appropriate interpretation of these changes in the context of physically active subjects.
PMCID: PMC3936967  PMID: 24627716
biomarkers; training; plasma volume; metabolism; cellular damage
2.  Interference of medical contrast media on laboratory testing 
Biochemia Medica  2014;24(1):80-88.
The use of contrast media such as organic iodine molecules and gadolinium contrast agents is commonplace in diagnostic imaging. Although there is widespread perception that side effects and drug interactions may be the leading problems caused by these compounds, various degrees of interference with some laboratory tests have been clearly demonstrated. Overall, the described interference for iodinate contrast media include inappropriate gel barrier formation in blood tubes, the appearance of abnormal peaks in capillary zone electrophoresis of serum proteins, and a positive bias in assessment of cardiac troponin I with one immunoassay. The interference for gadolinium contrast agents include negative bias in calcium assessment with ortho-cresolphthalein colorimetric assays and occasional positive bias using some Arsenazo reagents, negative bias in measurement of angiotensin converting enzyme (ACE) and zinc (colorimetric assay), as well as positive bias in creatinine (Jaffe reaction), total iron binding capacity (TIBC, ferrozine method), magnesium (calmagite reagent) and selenium (mass spectrometry) measurement. Interference has also been reported in assessment of serum indices, pulse oximetry and methaemoglobin in samples of patients receiving Patent Blue V. Under several circumstances the interference was absent from manufacturer-supplied information and limited to certain type of reagents and/or analytes, so that local verification may be advisable to establish whether or not the test in use may be biased. Since the elimination half-life of these compounds is typically lower than 2 h, blood collection after this period may be a safer alternative in patients who have received contrast media for diagnostic purposes.
PMCID: PMC3936969  PMID: 24627717
errors; analytical interference; contrast media; contrast agents; gadolinium
4.  A new device to relieve venipuncture pain can affect haematology test results 
Blood Transfusion  2014;12(Suppl 1):s6-s10.
In vitro diagnostic tests play a key role in patients’ management (e.g., guiding red blood cell transfusions). The aim of this study was to evaluate the impact of an innovative device (Buzzy®) which is claimed to be able to relieve venipuncture pain by means of cold and vibration. This device was applied during collection of venous blood by venipuncture for conventional haematology testing.
Materials and methods
Blood was drawn from 100 volunteers by a single, expert phlebotomist. A vein was located in the left forearm without applying a tourniquet but using a subcutaneous tissue transilluminator device, so that venous stasis was avoided. Blood samples were collected with a 20G straight needle directly into 4mL K3EDTA vacuum tubes. In sequence, external cold and vibration was established by Buzzy® on the right forearm −5 cm above the venipuncture site- for 1 minute before venipuncture and continued until the end of the same procedure already performed in the left forearm. Conventional haematological tests were performed using the same instrument (Sysmex® XE-2100D) in all cases.
When Buzzy® was applied before drawing blood, erythrocyte counts and associated parameters (i.e., haemoglobin and haematocrit) were higher, whereas platelet number, leucocyte count and differential were lower. Statistically and clinically significant differences (P <0.001) were observed for erythrocytes, haemoglobin and haematocrit.
From a practical perspective, cold-induced haemoconcentration promotes the efflux of water, diffusible ions and low molecular weight molecules from the vessel, thus increasing the concentration of other blood analytes at the puncture site. These variations may influence test results, especially for erythrocytes, haemoglobin and haematocrit. The novel Buzzy® device should, therefore, be used with caution when collecting blood for conventional haematological testing because of the observed bias introduced in some parameters.
PMCID: PMC3934256  PMID: 24120583
blood specimen collection; laboratory error; phlebotomy; preanalytical variability; reproducibility of results
5.  The effective reduction of tourniquet application time after minor modification of the CLSI H03-A6 blood collection procedure 
Biochemia Medica  2013;23(3):308-315.
The phlebotomists’ procedures are a still source of laboratory variability. The aim of this study was to verify the efficacy of minor modification in procedure for collection of diagnostic blood specimens by venipuncture from CLSI H03-A6 document is able to reduce the tourniquet application time.
Materials and methods:
Thirty phlebotomists were invited to participate. Each phlebotomist was trained individually to perform the new venipuncture procedure that shortens the time of tourniquet release and removal. The phlebotomy training program was delivered over 8h. After training, all phlebotomists were monitored for 20 working days, to guarantee the adoption of the correct new procedures for collection of diagnostic blood specimens. After this time frame the phlebotomists were evaluated to verify whether the new procedure for blood collection derived from CLSI H03-A6 document was effective to improve the quality process by decrease in tourniquet application time. We compared the tourniquet application time and qualitative difference of phlebotomy procedures between laboratories before and after phlebotomy training.
The overall mean ± SD tourniquet application time before and after this intervention were 118 ± 1 s and 30 ± 1 s respectively. Minor modifications in procedure for blood collection were able to reduce significantly the tourniquet application time (−88 s, P < 0.001).
The minor modifications in procedure for collection of diagnostic blood specimens by venipuncture from CLSI H03-A6 document were able to reduce the tourniquet application time. Now the proposed new procedure for collection of diagnostic blood specimens by venipuncture could be considered usefulness and should be put into practice by all quality laboratory managers and/or phlebotomy coordinators to avoid preanalytical errors regard venous stasis and guarantee patient safety.
PMCID: PMC3900072  PMID: 24266300
venous stasis; preanalytical phase; tourniquets; phlebotomy; practice guidelines
6.  Reduction of gross hemolysis in catheter-drawn blood using Greiner Holdex® tube holder 
Biochemia Medica  2013;23(3):303-307.
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.
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).
The use of Holdex for drawing blood from intravenous lines may be effective for reducing gross hemolysis.
PMCID: PMC3900079  PMID: 24266299
preanalytical variability; hemolysis; sample quality; intravenous line
7.  Influence of lean and fat mass on bone mineral density and on urinary stone risk factors in healthy women 
The role of body composition (lean mass and fat mass) on urine chemistries and bone quality is still debated. Our aim was therefore to determine the effect of lean mass and fat mass on urine composition and bone mineral density (BMD) in a cohort of healthy females.
Materials and methods
78 female volunteers (mean age 46 ± 6 years) were enrolled at the Stone Clinic of Parma University Hospital and subdued to 24-hour urine collection for lithogenic risk profile, DEXA, and 3-day dietary diary. We defined two mathematical indexes derived from body composition measurement (index of lean mass-ILM, and index of fat mass-IFM) and the cohort was split using the median value of each index, obtaining groups differing only for lean or fat mass. We then analyzed differences in urine composition, dietary intakes and BMD.
The women with high values of ILM had significantly higher excretion of creatinine (991 ± 194 vs 1138 ± 191 mg/day, p = 0.001), potassium (47 ± 13 vs 60 ± 18 mEq/day, p < 0.001), phosphorus (520 ± 174 vs 665 ± 186 mg/day, p < 0.001), magnesium (66 ± 20 vs 85 ± 26 mg/day, p < 0.001), citrate (620 ± 178 vs 807 ± 323 mg/day, p = 0.002) and oxalate (21 ± 7 vs 27 ± 11 mg/day, p = 0.015) and a significantly better BMD values in limbs than other women with low values of ILM. The women with high values of IFM had similar urine composition to other women with low values of IFM, but significantly better BMD in axial sites. No differences in dietary habits were found in both analyses.
Lean mass seems to significantly influence urine composition both in terms of lithogenesis promoters and inhibitors, while fat mass does not. Lean mass influences bone quality only in limb skeleton, while fat mass influences bone quality only in axial sites.
PMCID: PMC3853000  PMID: 24099643
Body composition; Lean mass; Fat mass; Bone mineral density; Urinary lithogenic risk factors
8.  Serum Oxidant and Antioxidant Status Following an All-Out 21-km Run in Adolescent Runners Undergoing Professional Training—A One-Year Prospective Trial 
This study investigated the 1-year longitudinal effect of professional training in adolescent runners on redox balance during intense endurance exercise. Changes in selected serum oxidant and antioxidant status in response to a 21-km running time trial in 10 runners (15.5 ± 1.3 years) undergoing professional training were evaluated twice in 12 months (pre- and post-evaluation). Venous blood samples were collected immediately before and 4-h following the 21-km run for analysis of serum concentrations of thiobarbituric acid-reactive substances (TBARS), xanthine oxidase (XO), catalase (CAT), reduced glutathione (GSH), superoxide dismutase (SOD), and total antioxidant capacity (T-AOC). In pre-evaluation trial, serum TBARS and SOD decreased after the 21-km run (p < 0.05) while XO, GSH, CAT and TAOC were unchanged. In post-evaluation trial, serum TBARS and SOD decreased, whereas XO and CAT increased post-exercise (p < 0.05). Furthermore, pre-exercise serum T-AOC, post-exercise serum XO, CAT, T-AOC (p < 0.05), and GSH (p = 0.057) appeared to be higher than the corresponding pre-evaluation values. The current findings suggest that a professional training regime in adolescent runners is not likely to jeopardize the development of their antioxidant defense. However, uncertainties in the maintenance of redox balance in runners facing increased exercise-induced oxidative stress as a consequence of training-induced enhancement of exercise capacity await further elucidation.
PMCID: PMC3742293  PMID: 23880864
adolescent; oxidative stress; antioxidant; endurance exercise
9.  Association between periodontal disease and Interleukin-1β +3953 and vitamin D receptor Taq1 genetic polymorphisms in an Italian caucasian population 
Annali di Stomatologia  2013;4(2):191-195.
Periodontal diseases entail a variety of conditions affecting the periodontium. The pathogenesis results from a complex interaction of genetic and environmental factors. Although there are evidences to confirm a role of genetic determinants, the outcome of the available studies is controversial and the largest part of the research has been carried out in Asian populations.
We investigated two polymorphisms in the genes encoding Interelukin-1β (IL-1β +3953 C>T; rs1143634) and vitamin D receptor (VDR Taq1; rs731236) in 42 Caucasian patients with chronic periodontal disease and 39 Caucasian subjects, matched for age and gender.
The IL-1β C allele was present in 100% of cases and 92% of controls (p=0.07), the T allele was present in 19% of cases and in 44% controls (p=0.017). The prevalence of the VDR Taq1 tt genotype was lower in patients as compared with controls (i.e., 10 versus 59%; p<0.01), whereas the tT and TT genotypes were disproportionally higher in patients than in cases (i.e., 62 versus 33% for tT and 29% versus 8% for TT; p<0.01).
The t allele was present in 71% of cases and 92% of controls (p=0.016), whereas the T allele was present in 90% of patients with periodontal disease and in 41% controls (p<0.01).
The results of this case control study at-test that the T allele of VDR Taq1 is strongly associated with periodontal disease, whereas the t allele of the IL-1β +3953 confers a slightly protection against the risk of periodontitis.
PMCID: PMC3755800  PMID: 23991270
periodontal diseases; periodontitis; genetics; interleukins; gene expression
10.  Incorrect order of draw could be mitigate the patient safety: a phlebotomy management case report 
Biochemia Medica  2013;23(2):218-223.
Procedures involving phlebotomy are critical for obtaining diagnostic blood specimens and represent a well known and recognized problem, probably among the most important issues in laboratory medicine. The aim of this report is to show spurious hyperkalemia and hypocalcemia due to inadequate phlebotomy procedure. The diagnostic blood specimens were collected from a male outpatient 45 years old, with no clinical complaints. The tubes drawing order were as follows: i) clot activator and gel separator (serum vacuum tube), ii) K3EDTA, iii) a needleless blood gas dedicated-syringe with 80 I.U. lithium heparin, directly connected to the vacuum tube holder system. The laboratory testing results from serum vacuum tube and dedicated syringe were 4.8 and 8.5 mmol/L for potassium, 2.36 and 1.48 mmol/L for total calcium, respectively. Moreover 0.15 mmol/L of free calcium was observed in dedicated syringe. A new blood collection was performed without K3EDTA tube. Different results were found for potassium (4.7 and 4.5 mmol/L) and total calcium (2.37 and 2.38 mmol/L) from serum vacuum tube and dedicated syringe, respectively. Also free calcium showed different concentration (1.21 mmol/L) in this new sample when compared with the first blood specimen. Based on this case we do not encourage the laboratory managers training the phlebotomists to insert the dedicated syringes in needle-holder system at the end of all vacuum tubes. To avoid double vein puncture the dedicated syringe for free calcium determination should be inserted immediately after serum tubes before EDTA vacuum tubes.
PMCID: PMC3900060  PMID: 23894868
clinical laboratory techniques; preanalytical phase; laboratory variability; blood specimen collection; EDTA contamination carryover
11.  Critical review and meta-analysis of spurious hemolysis in blood samples collected from intravenous catheters 
Biochemia Medica  2013;23(2):193-200.
A number of preanalytical activities strongly influence sample quality, especially those related to sample collection. Since blood drawing through intravenous catheters is reported as a potential source of erythrocyte injury, we performed a critical review and meta-analysis about the risk of catheter-related hemolysis.
Materials and methods:
We performed a systematic search on PubMed, Web of Science and Scopus to estimate the risk of spurious hemolysis in blood samples collected from intravenous catheters. A meta-analysis with calculation of Odds ratio (OR) and Relative risk (RR) along with 95% Confidence interval (95% CI) was carried out using random effect mode.
Fifteen articles including 17 studies were finally selected. The total number of patients was 14,796 in 13 studies assessing catheter and evacuated tubes versus straight needle and evacuated tubes, and 1251 in 4 studies assessing catheter and evacuated tubes versus catheter and manual aspiration. A significant risk of hemolysis was found in studies assessing catheter and evacuated tubes versus straight needle and evacuated tubes (random effect OR 3.4; 95% CI = 2.9–3.9 and random effect RR 1.07; 95% CI = 1.06–1.08), as well as in studies assessing catheter and evacuated tubes versus catheter and manual aspiration of blood (OR 3.7; 95% CI = 2.7–5.1 and RR 1.32; 95% CI = 1.24–1.40).
Sample collection through intravenous catheters is associated with significant higher risk of spurious hemolysis as compared with standard blood drawn by straight needle, and this risk is further amplified when intravenous catheter are associated with primary evacuated blood tubes as compared with manual aspiration.
PMCID: PMC3900066  PMID: 23894864
hemolysis; preanalytical variability; catheters; meta-analysis
12.  Avoidance to wipe alcohol before venipuncture is not a source of spurious hemolysis 
Biochemia Medica  2013;23(2):201-205.
It is still uncertain whether or not avoidance to let disinfectant alcohol dry at the site of venipuncture is a source of spurious hemolysis when drawing venous blood.
In a consecutive series of 52 outpatients referred for routine laboratory testing, venous blood was drawn by direct venipuncture with (odd group) or without (pair group) wiping 70% isopropyl alcohol at the site of venipuncture. A 3.5 mL evacuated tube with clot activator and gel separator was drawn from a vein of the upper limb, serum was immediately separated with standard centrifugation and tested for potassium, lactate dehydrogenase (LD), aspartate aminotransferase (AST) and hemolysis index (HI) on Roche Cobas.
No specimen was discarded for unsatisfactory venipuncture. No differences for age and gender were observed between groups. As regards the four parameters investigated, no significant differences could be observed between patients in whom blood was drawn with or without letting the alcohol dry. It is also noteworthy that no sample in both groups exceeded the conventional sample rejection threshold of cell-free hemoglobin.
The results of our prospective, randomized study attest that failure to wipe alcohol at the site of venipuncture should not be considered as a potential source of spurious hemolysis when drawing blood.
PMCID: PMC3900069  PMID: 23894865
preanalytical variability; hemolysis; alcohol; practice guideline
13.  Erythropoietin Receptor (EpoR) Agonism Is Used to Treat a Wide Range of Disease 
Molecular Medicine  2013;19(1):62-64.
The erythropoietin receptor (EpoR) was discovered and described in red blood cells (RBCs), stimulating its proliferation and survival. The target in humans for EpoR agonists drugs appears clear—to treat anemia. However, there is evidence of the pleitropic actions of erythropoietin (Epo). For that reason, rhEpo therapy was suggested as a reliable approach for treating a broad range of pathologies, including heart and cardiovascular diseases, neurodegenerative disorders (Parkinson’s and Alzheimer’s disease), spinal cord injury, stroke, diabetic retinopathy and rare diseases (Friedreich ataxia). Unfortunately, the side effects of rhEpo are also evident. A new generation of nonhematopoietic EpoR agonists drugs (asialoEpo, Cepo and ARA 290) have been investigated and further developed. These EpoR agonists, without the erythropoietic activity of Epo, while preserving its tissue-protective properties, will provide better outcomes in ongoing clinical trials. Nonhematopoietic EpoR agonists represent safer and more effective surrogates for the treatment of several diseases such as brain and peripheral nerve injury, diabetic complications, renal ischemia, rare diseases, myocardial infarction, chronic heart disease and others.
PMCID: PMC3646093  PMID: 23615965
14.  Ex vivo erythrocyte generation and blood doping 
Blood Transfusion  2013;11(2):161-163.
PMCID: PMC3626461  PMID: 23114519
15.  Blood Phosphorus and Magnesium Levels in 130 Elite Track and Field Athletes 
This study tested the clinical utility and relevance of serum phosphorus and magnesium as markers possibly useful to monitor training in athletes.
Phosphorus and magnesium serum concentrations of 130 elite track and field athletes (65 males and 65 females, age range 20-30 years) from the National Athletics Sports Medicine Center database in Thessaloniki, Greece were measured.
Abnormal results were found in 61 (47%) athletes (32 men and 29 women). In male athletes, serum phosphate was higher than normal in 18% and decreased in 1.5%, whereas serum magnesium concentration was higher in 26%, and lower in 3%. Regarding female athletes, higher serum phosphate and magnesium levels were detected in 26% and 17% respectively, whereas decreased serum magnesium was found in 3%. The most common alterations were higher serum phosphate (29/61, 47%) and magnesium concentrations (28/61, 46%). Abnormalities of serum phosphorus and magnesium concentrations were detected in almost half of the athletes. Hyperphosphataemia and hypermagnesaemia were the most common abnormalities.
The reference intervals used for general population cannot be used for athletes. Given the lack of pathological manifestations, the physiopathological significance of these findings is uncertain. Further studies on the interpretation of reported ion concentrations in athletes should take in account the type of sport practiced and also the possible variations during the training and competition season.
PMCID: PMC3685160  PMID: 23785576
Athletes; Hyperphosphataemia; Hypophosphatemia; Hypomagnesaemia; Hypermagnesaemia
16.  Glycogen phosphorylase isoenzyme BB in the diagnosis of acute myocardial infarction: a meta-analysis 
Biochemia Medica  2013;23(1):78-82.
Early diagnosis is crucial for management of patients with suspected acute myocardial infarction (AMI). Among innovative and promising biomarkers, the recent interest raised on glycogen phosphorylase isoenzyme BB (GPBB) has prompted us to perform a meta-analysis of published studies.
Materials and methods:
A systematic electronic search was carried out on PubMed, Web of Science and Google Scholar, with no date restriction, to retrieve all articles that have investigated the early diagnostic performance of GPBB in patients with suspected AMI, and directly reported or allowed calculation of sensitivity and specificity. A meta-analysis of the reported sensitivity and specificity of each study and pooled area under the curve (AUC) was then performed by random effect approach. Heterogeneity was assessed by I-square statistics.
Eight studies were finally selected for analysis (941 subjects; 506 cases and 435 controls), with a high heterogeneity (I-squared, 86.3%). The resulting pooled estimates and 95% confidence interval were 0.854 (0.801–0.891) for sensitivity, 0.767 (0.713–0.815) for specificity, 0.826 (0.774–0.870) for negative predictive value, 0.802 (0.754–0.844) for positive predictive value, and 0.754 (0.602–0.907) for AUC. In those studies that have simultaneously assessed GPBB and a troponin immunoassay, the combination of these biomarkers did not significantly improve the performance of troponin alone.
GPBB does not meet the current requirements for an efficient diagnosis of AMI when used as a stand-alone test, whereas its combination with troponin merits further investigation in larger trials.
PMCID: PMC3900091  PMID: 23457768
myocardial infarction; ischemic heart disease; glycogen phosphorylase isoenzyme BB; GPBB
17.  Evaluation of sample hemolysis in blood collected by S-Monovette® using vacuum or aspiration mode 
Biochemia Medica  2013;23(1):64-69.
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.
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.
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.
PMCID: PMC3900093  PMID: 23457766
hemolysis; hemolyzed specimens; preanalytical variability; blood collection tube; blood drawing
19.  Impact of the phlebotomy training based on CLSI/NCCLS H03-A6 - procedures for the collection of diagnostic blood specimens by venipuncture. 
Biochemia Medica  2012;22(3):342-351.
The activities involving phlebotomy, a critical task for obtaining diagnostic blood samples, are poorly studied as regards the major sources of errors and the procedures related to laboratory quality control. The aim of this study was to verify the compliance with CLSI documents of clinical laboratories from South America and to assess whether teaching phlebotomists to follow the exact procedure for blood collection by venipuncture from CLSI/NCCLS H03-A6 - Procedures for the Collection of Diagnostic Blood Specimens by Venipuncture might improve the quality of the process.
Materials and methods:
A survey was sent by mail to 3674 laboratories from South America to verify the use of CLSI documents. Thirty skilled phlebotomists were trained with the CLSI H03-A6 document to perform venipuncture procedures for a period of 20 consecutive working days. The overall performances of the phlebotomists were further compared before and after the training program.
2622 from 2781 laboratories that did answer our survey used CLSI documents to standardize their procedures and process. The phlebotomists’ training for 20 days before our evaluation completely eliminated non-conformity procedures for: i) incorrect friction of the forearm, during the cleaning of the venipuncture site to ease vein location; ii) incorrect sequence of vacuum tubes collection; and iii) inadequate mixing of the blood in primary vacuum tubes containing anticoagulants or clot activators. Unfortunately the CLSI H03-A6 document does not caution against both unsuitable tourniquet application time (i.e., for more than one minute) and inappropriate request to clench the fist repeatedly. These inadequate procedures were observed for all phlebotomists.
We showed that strict observance of the CLSI H03-A6 document can remarkably improve quality, although the various steps for collecting diagnostic blood specimens are not a gold standard, since they may still permit errors. Tourniquet application time and forearm clench should be verified by all quality laboratory managers in the services. Moreover, the procedure for collecting blood specimens should be revised to eliminate this source of laboratory variability and safeguard the quality.
PMCID: PMC3900046  PMID: 23092065
phlebotomy; blood specimen collection; tourniquet application time; CLSI documents; pre-analytic variability; extra- analytical variability
20.  The new oral anticoagulants and the future of haemostasis laboratory testing 
Biochemia Medica  2012;22(3):329-341.
The tests currently employed within most haemostasis laboratories to monitor anticoagulant therapy largely comprise the prothrombin time (PT)/ International Normalised Ratio (INR) and the activated partial thromboplastin time (APTT). These are respectively used to monitor Vitamin K antagonists (VKAs) such as warfarin, and unfractionated heparin. Additional tests that laboratories may also employ for assessing or monitoring unfractionated heparin include thrombin time (TT) and the anti-Xa assay, which can also be used to monitor low molecular weight heparin. Several new anti-thrombotic agents have recently emerged, or are in the final process of clinical evaluation. These novel drugs that include Dabigatran etexilate and Rivaroxaban would not theoretically require monitoring; however, testing is useful in specific situations. The tests currently used to monitor VKAs and heparin are typically either too sensitive or too insensitive to the new drugs to be used as ‘typically performed in laboratories’, and may thus require some methodological adjustments to increase or decrease their sensitivity. Alternately, different tests may be better employed in these assessments. Whatever the case, laboratories may soon be performing a reduced or possibly increased number of tests, the same kind of tests but perhaps differently, or conceivably different assay panels. Specific laboratory guidance on the choice of the appropriate test to be ordered according to the drug being administered, as well as on appropriate interpretation of test results, will also be necessary. The current report reviews the current state of play and provides a glimpse to the possible future of the coagulation laboratory.
PMCID: PMC3900050  PMID: 23092064
haemostasis; coagulation; laboratory tests; anticoagulants; anti-thrombotics, Dabigatran, Rivaroxaban
21.  K3EDTA Vacuum Tubes Validation for Routine Hematological Testing 
ISRN Hematology  2012;2012:875357.
Background and Objective. Some in vitro diagnostic devices (e.g, blood collection vacuum tubes and syringes for blood analyses) are not validated before the quality laboratory managers decide to start using or to change the brand. Frequently, the laboratory or hospital managers select the vacuum tubes for blood collection based on cost considerations or on relevance of a brand. The aim of this study was to validate two dry K3EDTA vacuum tubes of different brands for routine hematological testing. Methods. Blood specimens from 100 volunteers in two different K3EDTA vacuum tubes were collected by a single, expert phlebotomist. The routine hematological testing was done on Advia 2120i hematology system. The significance of the differences between samples was assessed by paired Student's t-test after checking for normality. The level of statistical significance was set at P < 0.05. Results and Conclusions. Different brand's tubes evaluated can represent a clinically relevant source of variations only on mean platelet volume (MPV) and platelet distribution width (PDW). Basically, our validation will permit the laboratory or hospital managers to select the brand's vacuum tubes validated according to him/her technical or economical reasons for routine hematological tests.
PMCID: PMC3409519  PMID: 22888448
22.  Influence of a Regular, Standardized Meal on Clinical Chemistry Analytes 
Annals of Laboratory Medicine  2012;32(4):250-256.
Preanalytical variability, including biological variability and patient preparation, is an important source of variability in laboratory testing. In this study, we assessed whether a regular light meal might bias the results of routine clinical chemistry testing.
We studied 17 healthy volunteers who consumed light meals containing a standardized amount of carbohydrates, proteins, and lipids. We collected blood for routine clinical chemistry tests before the meal and 1, 2, and 4 hr thereafter.
One hour after the meal, triglycerides (TG), albumin (ALB), uric acid (UA), phosphatase (ALP), Ca, Fe, and Na levels significantly increased, whereas blood urea nitrogen (BUN) and P levels decreased. TG, ALB, Ca, Na, P, and total protein (TP) levels varied significantly. Two hours after the meal, TG, ALB, Ca, Fe, and Na levels remained significantly high, whereas BUN, P, UA, and total bilirubin (BT) levels decreased. Clinically significant variations were recorded for TG, ALB, ALT, Ca, Fe, Na, P, BT, and direct bilirubin (BD) levels. Four hours after the meal, TG, ALB, Ca, Fe, Na, lactate dehydrogenase (LDH), P, Mg, and K levels significantly increased, whereas UA and BT levels decreased. Clinically significant variations were observed for TG, ALB, ALT, Ca, Na, Mg, K, C-reactive protein (CRP), AST, UA, and BT levels.
A significant variation in the clinical chemistry parameters after a regular meal shows that fasting time needs to be carefully considered when performing tests to prevent spurious results and reduce laboratory errors, especially in an emergency setting.
PMCID: PMC3384805  PMID: 22779065
Blood specimen collection; Clinical laboratory techniques; Diagnostic errors; Eating; Fasting; Postprandial period; Reference values; Reproducibility of results; Quality control; Specimen handling
23.  Foot-strike haemolysis after a 60-km ultramarathon 
Blood Transfusion  2012;10(3):377-383.
The various contributors to sport-related anaemia include increased plasma volume, exercise-induced oxidative stress, increased body temperature, acidosis, gastrointestinal bleeding, acute and chronic inflammation as well as compression and damage of red blood cells (RBC) in the capillaries within the contracting muscles. The effective contribution of foot-strike haemolysis is unclear.
Materials and methods.
We studied 18 Caucasian male athletes (mean age, 42 years; range, 34–52 years) before and immediately after a 60-km ultramarathon. Laboratory investigations included the haematological profile along with haptoglobin, potassium, aspartate aminotransferase (AST), creatine kinase (CK), lactate dehydrogenase (LDH) and albumin concentrations and a haemolysis index (HI).
No significant variations were found in post-exercise values of haemoglobin, RBC count and haematocrit. Mean corpuscular volume and haptoglobin were significantly decreased, whereas RBC distribution width was increased. The concentration of haptoglobin was reduced by approximately 50%, whereas enzyme concentrations were all remarkably increased. The HI remained below 0.5 g/L. After adjusting for plasma volume change, the increases were 1.7% for potassium (P=0.17), 30% for AST (P<0.01), 49% for LDH (P<0.01) and 2.39-fold for CK (P<0.01). A statistically significant association was found between haemoconcentration-adjusted variations of CK and those of AST (r=0.803; P<0.01) and LDH (r=0.551; P=0.02).
This is the first study demonstrating that long-distance running does not induce clinically significant changes in haemoglobin, haematocrit, RBC count or potassium concentration. The significant post-exercise decrease of haptoglobin reflects a certain degree of haemolysis, but the concentration of cell-free haemoglobin remaining below 0.5 g/L and the non-significant variation in RBC count both indicate that the foot-strike haemolysis is very modest or even clinically negligible.
PMCID: PMC3417738  PMID: 22682343
sport; physical exercise; marathon; haemolysis
24.  Serum Oxidant and Antioxidant Status in Adolescents Undergoing Professional Endurance Sports Training 
This study evaluated the impact of professional training on serum oxidant and antioxidant status in adolescent endurance athletes and compared it with that of untrained individuals. Firstly, serum thiobarbituric-acid-reactive substances (TBARSs), xanthine oxidase (XO), catalase (CAT), reduced glutathione (GSH), superoxide dismutase (SOD), and total antioxidant capacity (T-AOC) were measured in 67 male runners, cyclists, and untrained adolescents. Seven-day dietary intakes were also assessed. Secondly, for age- and Tanner-stage-matched comparison, 36 out of the 67 subjects (12 for each group) were then selected and investigated. In cyclists, XO, GSH, and CAT were higher as compared with runners and controls. The CAT in runners, but not GSH and XO, was also higher than in controls. TBARS, T-AOC, and SOD did not differ among the study populations. Regarding the inter-individual relationships among serum redox statuses and dietary nutrient intakes, significant correlations were noted in CAT versus carbohydrates, protein, magnesium, and manganese; GSH versus carbohydrates, protein, fat, selenium, zinc, iron, and magnesium; XO versus cholesterol; CAT versus GSH. These findings suggest that the resting blood redox balance in the professional adolescent athletes was well maintained partly by the increase of individual antioxidant in adaptation to chronic exercise.
PMCID: PMC3345234  PMID: 22577491

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