Plain Glass Tubes Research Articles

Investigation of the effects of storage with preservatives at room temperature or refrigeration without preservatives on urinalysis results for samples from healthy dogs

To compare urinalysis results for canine urine samples stored in preservative-containing tubes at room temperature (20°C to 25°C [68°F to 77°F]) or refrigerated at 4°C (39.2°F) in plain glass tubes with results for the same samples immediately after collection. Urine samples from 20 healthy dogs. Urine samples (1/dog) were divided into 6 aliquots (3 in preservative-containing tubes and 3 in plain glass tubes). Preservative-containing tubes were stored at room temperature and plain glass tubes were refrigerated. Urinalysis was performed 0, 24, and 72 hours after collection. Results for both storage conditions were compared with results for a reference sample (the 0-hour [immediate post-collection] aliquot in a plain glass tube) by Spearman correlation analysis with pairwise tests for selected variables. Physical variables (urine color and turbidity with and without centrifugation) for both storage conditions had high (rs = 0.7 to 0.9) or very high (rs = 0.9 to 1.0) degrees of positive correlation with reference sample results at all time points, except for color at 24 hours. Similar results were found for all biochemical variables with storage up to 72 hours. For microscopic characteristics, correlation with reference sample results ranged from low or nonsignificant to very high under both storage conditions. Results suggested that if a delay in urinalysis is expected, use of the preservative-containing tubes evaluated in this study may be a viable option for sample storage. Further research is warranted to assess direct comparability of results to those of freshly collected samples and use of these tubes to store samples from dogs with conditions affecting the urinary tract.

Concentrated Growth Factor Matrices Prepared Using Silica-Coated Plastic Tubes Are Distinguishable From Those Prepared Using Glass Tubes in Platelet Distribution: Application of a Novel Near-Infrared Imaging-Based, Quantitative Technique

Platelet-rich fibrin (PRF) matrices were originally prepared using plain glass tubes without the aid of coagulation factors because coagulation factor XII is activated by glass surfaces. Recently, the use of silica-coated plastic tubes as a substitute of glass tubes has been recommended for PRF preparation. This recommendation is owing not only to the shortage of glass tubes for medical use in the market, but also the higher coagulation activity of silica-coated plastic tubes and equal quality of PRF. However, these matrices are not the same. To evaluate the differences, we compared glass- and silica-coated plastic tubes in terms of platelet distribution and quantity in concentrated growth factors (CGF). CGF matrices were immediately prepared from freshly collected blood samples, fixed after red thrombus removal, and divided into two equal pieces sagittally. One piece was used for CD41 detection and the other was applied as an isotype control. Platelet distribution in CGF matrices was examined, without embedding or sectioning, by a novel method using invisible near-infrared imaging. The dehydrated membranous CGF matrix was more transparent. Thus, the fluorescence signal was clearly detectable with less scattering. Platelets were distributed mainly in the distal side of the glass-prepared CGF matrix, but homogeneously in the silica-prepared CGF matrix. Platelet count was positively correlated with fluorescence intensity. Although not yet fully developed, this imaging technique enabled us to recognize the differences in platelet distribution and quantity in CGF matrices by excluding bias caused by the technical limitations of scanning electron microscopy and conventional immunohistochemical methods.

Striking Differences in Platelet Distribution between Advanced-Platelet-Rich Fibrin and Concentrated Growth Factors: Effects of Silica-Containing Plastic Tubes.

Compared with platelet-rich plasma, the preparation of platelet-rich fibrin (PRF) is simple and has not been overly modified. However, it was recently demonstrated that centrifugation conditions influence the composition of PRF and that silica microparticles from silica-coated plastic tubes can enter the PRF matrix. These factors may also modify platelet distribution. To examine these possibilities, we prepared PRF matrices using various types of blood-collection tubes (plain glass tubes and silica-containing plastic tubes) and different centrifugation speeds. The protocols of concentrated growth factors and advanced-PRF represented high- and low-speed centrifugation, respectively. Platelet distribution in the PRF matrix was examined immunohistochemically. Using low-speed centrifugation, platelets were distributed homogeneously within the PRF matrix regardless of tube types. In high-speed centrifugation, platelets were distributed mainly on one surface region of the PRF matrix in glass tubes, whereas in silica-coated tubes, platelet distribution was commonly more diffusive than in glass tubes. Therefore, both blood-collection tube types and centrifugal conditions appeared to influence platelet distribution in the PRF matrix. Platelets distributed in the deep regions of the PRF matrix may contribute to better growth factor retention and release. However, clinicians should be careful in using silica-coated tubes because their silica microparticles may be a health hazard.

Evidence for Contamination of Silica Microparticles in Advanced Platelet-Rich Fibrin Matrices Prepared Using Silica-Coated Plastic Tubes.

Platelet-rich fibrin (PRF) therapy has been widely applied in regenerative dentistry, and PRF preparation has been optimized to efficiently form fibrin clots using plain glass tubes. Currently, a shortage of commercially available glass tubes has forced PRF users to utilize silica-coated plastic tubes. However, most plastic tubes are approved by regulatory authorities only for diagnostic use and remain to be approved for PRF therapy. To clarify this issue, we quantified silica microparticles incorporated into the PRF matrix. Blood samples were collected into three different brands of silica-containing plastic tubes and were immediately centrifuged following the protocol for advanced-PRF (A-PRF). Advanced-PRF-like matrices were examined using a scanning electron microscope (SEM), and silica microparticles were quantified using a spectrophotometer. Each brand used silica microparticles of specific size and appearance. Regardless of tube brands and individual donors, significant, but not accidental, levels of silica microparticles were found to be incorporated into the A-PRF-like matrix, which will be consequently incorporated into the implantation sites. Presently, from the increasing data for cytotoxicity of amorphous silica, we cannot exclude the possibility that such A-PRF-like matrices negatively influence tissue regeneration through induction of inflammation. Further investigation should be performed to clarify such potential risks.

Selective amelioration of chronic resraint stress effects on il-6, liver function and erythrocyte indices by the chemical alprazolam drug or the herb valerian extract in rats

The present study aimed at evaluating the ameliorative efficacy of daily oral treatment of adult male rats with either the chemical alprazolam (ALZ) drug (0.3 mg/kg b.wt) or the herb valerian (VAL) extract (300 mg/kg b.wt) against changes in serum levels of IL-6 and liver function biomarkers ALT, AST , ALP activity . and erythrocyte indices induced by daily (chronic) exposure of rats to restraint (RST) stress (2.5 h /day) for 4 weeks . Rats were divided into 6 groups (10 rats each) ; group 1 : Non-RST control group; group 2 : RST rats ; group 3 : non-RST + ALZ treated rats ; group 4 : non- RST + VAL extract treated rats ; group 5 : RST + ALZ treated rats; group 6 : RST + VAL extract treated rats . Blood samples were withdrawn from the retro- orbital venous plexus of each rat under slight ether anesthesiainto plain and EDTA glass tubes for separating sera samples and measuring erythrocyte indices respectively. RST exposed rats showed significant increases in serum levels of IL-6 , ALT, AST , ALP activity and significant increases in RBCs count, values of Hb, HCT, MCV, MCH and MCHC. Administration of ALZ alone to non-RST rats had no significant effect on serum level of IL-6 , but showed significant increase in serum activity of ALT, AST , ALP, and significant decreases in total RBCs counts , Hb , HCT, MCV, MCH and MCHC values. Whereas administration of VAL extract alone to non-RST rats did not cause any significant change in all former parameters . ALZ treatment of RST rats selectively did not ameliorate stress induced changes in ALT, AST , ALP activity, but markedly protected erythrocyte indices against RST stress effects . On the contrary , administration of VAL extract to RST rats revealed its ability to markedly and selectively ameliorate the changes induced by RST stress in serum IL-6 level and the activity of liver enzymes measured , but had weaker ameliorative effect on stress induced changes in erythrocyte indices measured.

Vascular function and brain-derived neurotrophic factor: The functional capacity factor.

Brain-derived neurotrophic factor (BDNF) is essential for neurocognitive function. This study aims at establishing a plausible link between level of serum BDNF, functional capacity (FC), and vascular function in 181 young (age 25.5±9.1 years old), apparently healthy adults. Fasting blood samples were drawn from participants' antecubital veins into plain glass tubes while they were in a sitting position to evaluate serum BDNF using enzyme-linked immunosorbent assay (ELISA). Mercury-in-silastic strain-gauge plethysmography was used to determine arterial function indices, blood flow and vascular resistance at rest and following 5 minutes of arterial ischemia. The 6-minute walk distance (6MWD) test was used to determine FC, according to the American Thoracic Society Committee on Proficiency Standards for Clinical Pulmonary Function Laboratories guidelines. It was conducted in an enclosed corridor on a flat surface with a circular track 33 meters long. The walking course was demarcated with bright colored cones. The 6MWD correlated with BDNF (r=0.3, p=0.000), as well as with forearm blood inflow (r=0.5, p=0.000) and vascular resistance (r = -0.4, p=0.000). Subsequent comparison showed that BDNF and blood inflow were greater (p<0.05) while vascular resistance was less (p<0.05) in participants who achieved a longer 6MWD. Similarly, BDNF correlated with forearm blood inflow (r=0.4, p=0.000) and vascular resistance (r = -0.4, p=0.000). Subsequent comparison showed improved vascular function (p<0.05) in the participants with greater BDNF. In conclusion, these findings might suggest that improved vascular function in individuals with greater FC is mediated, at least partially, by an enhanced serum BDNF level.

Stability of oleander extract and oleandrin in sera stored in plastic serum separator tubes

Oleander poisoning is common in the southern parts of the United States as well as part of Europe and South East Asia. The fluorescence polarization immunoassay (FPIA) for digoxin cross-reacts with oleandrin, the major poison of oleander extract and can be used for rapid detection of oleander poisoning. However, for medicolegal investigation, the presence of oleandrin should be confirmed by a more sophisticated analytical technique such as liquid chromatography combined with tandem mass spectrometry. We studied stability of oleander extract as well as oleandrin in serum samples stored in Becton Dickinson plastic serum separator tubes (SSTs) by comparing values obtained in parallel specimens stored in plain glass tubes and in red-topped plastic tubes. Oleander leaves were extracted using absolute ethanol, and drug-free serum pools were supplemented with oleander extract or ethanolic solution of pure oleandrin. Aliquots of the supplemented pools were then stored in plain glass tubes, red-topped plastic tubes or SSTs and the stability of oleander extract or oleandrin in sera were studied by the FPIA digoxin assay for varying times up to seven days. We observed no statistically significant change in apparent digoxin concentrations in specimens stored in SSTs even after seven days of storage compared to original values observed in the supplemented serum pools. Specimens can be stored in SSTs for up to seven days prior to analysis of oleandrin concentration.

Soluble CD40 Ligand Measurement Inaccuracies Attributable to Specimen Type, Processing Time, and ELISA Method

CD40 ligand (CD40L) is a member of the tumor necrosis factor superfamily and is produced in a variety of cells, including platelets. The soluble form (sCD40L) is a mediator of both inflammatory and hemostasis processes and has been implicated in the pathogenesis of atherosclerosis. Clinical studies have revealed increased sCD40L in patients with unstable angina (1) and identified an association between increased sCD40L and future risk for death or nonfatal myocardial infarction (2)(3). While prospectively measuring sCD40L in a cohort of persons at risk for cardiovascular complications, we identified both preanalytical and analytical sources of error. This report documents the effects of specimen type (serum and plasma), processing (time and temperature), and commercial reagent selection on sCD40L ELISA results. These findings raise concerns about the accuracy of sCD40L results reported in recent clinical studies. After obtaining informed consent, we enrolled 147 patients older than 60 years referred for diagnostic cardiac catheterization in an Institutional Review Board-approved study to evaluate the value of clinical, echocardiographic, and biomarker variables for prediction of future cardiovascular complications. When combined with clinical predictors, B-type natriuretic peptide and C-reactive protein, but not sCD40L, were independent predictors of death or cardiovascular hospitalization at 6 months (data not shown). The unexpectedly poor correlation between undiluted plasma sCD40L results and clinical outcomes in this study motivated us to perform the following investigations. Venous blood from the 147 study participants [mean (SD) age, 70.8 (6.9) years] was collected into plastic tubes containing tripotassium EDTA (BD Vacutainer; Becton Dickinson) before catheterization and placed on ice for 1–4 h before processing. Blood from 10 control individuals [mean (SD) age, 38.7 (8.4) years] was collected into both EDTA and plain glass tubes (Becton Dickinson) and maintained at room temperature for 30 min before processing. Study and control samples were centrifuged twice: …

Variations in Pleural Fluid WBC Count and Differential Counts With Different Sample Containers and Different Methods

To compare the results of pleural fluid analysis for WBC counts and differential cell counts as follows: (1) counting performed manually vs that performed by an automated cell counter; (2) cells collected in different types of specimen containers; and (3) cell counts performed at 4 and 24 h postthoracentesis. Twenty-eight pleural fluid samples were each collected in five different containers (ie, ethylenediaminetetraacetic acid (EDTA)-treated glass, citrate-treated glass, heparinized glass, plain glass, and plain plastic tubes). The WBC counts and differential cell counts were obtained manually (on the EDTA tube) and with an automated counter on all tubes within 4 h of collection, and again after 24 h of refrigeration. There was a close correlation between the WBC counts obtained manually and those obtained with the automated counter from the pleural fluid samples collected in the EDTA tubes (r = 0.92). With the automated counter, the pleural fluid WBC counts were similar among the three tubes containing anticoagulants, but the counts obtained from the tubes without anticoagulants were significantly lower. The differential cell counts obtained manually and those obtained with the automated cell counter differed substantially. Although the percentage of lymphocytes was similar, the automated counter was inaccurate in differentiating neutrophils from large monocytes or mesothelial cells. The WBC counts obtained within 4 h of collection and 24 h after collection were virtually identical. The WBC counts obtained manually and with the automated counter from pleural fluid samples in EDTA tubes correlated very closely. The pleural fluid WBC count was lower if the pleural fluids had been collected in tubes without an anticoagulant. Automated WBC counts from pleural fluid specimens were inaccurate, possibly due to difficulty in separating neutrophils from monocyte/mesothelial cells. Refrigerated storage for up to 24 h had no significant effect on the total WBC count or on the WBC count differential regardless of the tube utilized.

Effect of a protease inhibitor on in vitro stability of intact parathyroid hormone.

We investigated whether increased protease activity explains the increased in vitro degradation of intact parathyroid hormone (iPTH) observed in serum when compared to EDTA plasma. Pre-dialysis blood samples for iPTH were taken from 11 patients with chronic renal failure and collected into plain glass tubes, tubes containing 200 KIU/mL aprotinin (a protease inhibitor) and EDTA tubes. All sample aliquots were separated at 20 min, 1 h, 2 h, 4 h, 8 h and 24 h post collection. Over 24 h, iPTH concentrations remained unchanged in EDTA tubes. iPTH concentrations were significantly lower in both plain tubes (P < 0.01) and aprotinin tubes (P < 0.001) at 24 h when compared to the baseline sample (20 min). At 24 h, iPTH concentrations in EDTA tubes were higher than in plain tubes (P < 0.001) and aprotinin tubes (P < 0.01). The addition of aprotinin to plain tubes significantly reduced the degradation of iPTH (P < 0.05) at 24 h. Aprotinin significantly reduces the in vitro degradation of iPTH in plain tubes at 24 h from 24.7% to 9.6%. We suggest that increased protease activity contributes to the decline in serum iPTH over time. As this is observed in serum and not plasma it suggests that the increased protease activity may be due to the clotting process.

INVESTIGATION OF ANTITHROMBOGENICITY OF MAGNETITE

In examining the antithrombogenicity of magnetite, we investigated the interaction between platelets and magnetite, Methods 1) The coagulation onset times (COTS) of platelet-rich plasma (PRP) anti platelet-free plasma (PFP) in glass tubes coated with magnetite were determined with a rheometer, which yields COTs of 20 min and more for materials considered antithrombogenic. 2) Platelet activation was investigated by scanning electron microscopy (SEM) 20 min after PRP, in the absence of calcium, was plated on magnetite and segmented polyurethane (SPU) sheets (N = 4). Results 1) The COTs of PRP in 11 magnetite and plain glass tubes were 27.1 ± 8.9 and 2.3 ± 0.6 min, respectively (p < 0.0001). 2) The COTS of PFP in 4 glass tubes were 5.5 ± 1.0 min, hut the PFP experiment in 4 magnetite tubes was stopped at 140 min because PFP did not coagulate at all. 3) SEM study showed that hardly any platelets adhered to the magnetite and SPU sheets, covering only 0.55 ± 0.14% and 0.60 ± 0.30%, respectively, of the entire surface area (NS). Approximately 60 ± 70% of the adhering platelets protruded pseudo-pods. Conclusions Our findings indicate that magnetite does not activate the clotting factors of the contact phase at all, but may activate platelets to a very small extent. In conclusion, these findings indicate that magnetite has excellent antithrombogenicity.

Inhibition of fibrin monomer polymerisation by myeloma immunoglobulin.

Myelomatosis was diagnosed in a 64 year old man on the basis of a serum paraprotein band (type IgG lambda, 42 g/l), plasma cell infiltration of bone marrow, and multiple lytic lesions evident on skull x ray picture. Blood specimens taken into plain glass tubes showed bulky gelatinous clot formation with minimal clot retraction. Coagulation tests were significantly abnormal with an increase in thrombin time, prothrombin time, and reptilase time. The possibility that the paraprotein was interfering with fibrin production was investigated. The rate of fibrin monomer polymerisation (measured turbidometrically) was reduced in patient plasma compared with control plasma. Although purified fibrin monomer prepared from the patient's fibrinogen polymerised normally, the addition of purified paraprotein caused a dose dependent reduction in the rate of polymerisation. These results suggest that the paraprotein was impairing fibrin formation by inhibiting fibrin monomer polymerisation. After chemotherapy the paraprotein concentration decreased and the coagulation results returned to normal.

Changes in coagulation and fibrinolysis in horses during exercise

SUMMARY Changes in clotting time (ct) and fibrinolytic actvity (fa) were evaluated in 6 mature, female horses during exercise. Two trials were performed on consecutive days, using a randomized crossover design. Each mare was assigned to either an exercise trial or a control trial on the first day, and to the alternate trial 24 hours later. Mares exercised for 20 minutes on a treadmill at an elevation of 2° and a velocity of 5 m/s. Venous blood samples were collected immediately before exercise, at 4, 8, 12, 16, and 20 minutes during exercise, and 15 minutes after cessation of exercise. Blood was placed into plain glass tubes for determination of CT, and into chilled, citrated tubes for determination of FA, plasminogen/plasmin complex activity (plg), one-stage prothrombin time (ospt), activated partial thromboplastin time (aptt), and antithrombin-III (at-III) activity. There were significant differences (P &lt; 0.05) between the control and exercise groups for ct, fa, and plg. During exercise, clotting time decreased from 21.5 ± 1.6 minutes to 9.9 ± 1.6 minutes (mean ± sd; P &lt; 0.05), without significant changes in ospt, aptt, or at-III. Fibrinolytic activity and plg increased (P &lt; 0.05) during exercise. Changes in ct, fa, and plg were significant at 4 minutes of exercise, remained altered until the end of exercise, and returned to baseline values by 45 minutes of recovery. Clotting time, ospt, aptt, fa, at-III, and plg did not change (P &gt; 0.05) during control trials.

Actual ionized calcium and pH in blood collected in capillary or evacuated tubes.

Results are presented for the concentration of actual ionized calcium (cCa2+), the actual pH and ionized calcium at pH = 7.4 [cCa2+(7.4)] measured in capillary whole blood, venous serum from vacuum tubes (Venoject) and serum equilibrated to pCO2 approximately equal to 5.3 kPa from 57 renal patients. The mean value for cCa2+ was higher for venous serum than for capillary blood (1.29 mmol/l versus 1.26 mmol/l), corresponding to the difference in pH (in venous serum). This difference could be eliminated by separating the serum at 4 degrees C rather than room temperature. The cCa2+ (7.4) was the same for capillary blood, venous serum (Venoject) and equilibrated serum from blood collected in plain glass tubes, Venous serum could be stored at 4 degrees C for 4 h without significant change in cCa2+ and pH, and for 1 week at 4 degrees C for the measurement of cCa2+(7.4). In patients with acid-base disturbance cCa2+ is preferable to the measurement of cCa2+(7.4).

Increase in Serum Testosterone following Contact with Blood Cells

Whilst validating a testosterone assay, the normal range for females was found to be substantially higher than that derived when the original method was developed (M J Wheeler, St Thomas' Hospital, London). There was an excellent agreement between the results obtained from 13 samples with serum testoster­ one in the range 0·8-4·0 nmol/L, which were analysed by both laboratories (r=0·992, n=13). This indicated that the higher normal range was not due to an assay bias. Subsequent investiga­ tions, described here, strongly suggest that the higher normal range results from a difference in specimen handling. Blood was collected from eight females attending a subfertility clinic and from five female laboratory staff. Each blood sample was divided between two plain glass tubes and, on receipt in the laboratory, the serum from one tube was separated and stored at - 20°C until analysis. The other tube was left at room temperature (2o-25°C) for between 1 and 5 days before separation of serum and storage at -200C. Testosterone was measured by radioimmu­ noassay following ether extraction of the

Prostanoid synthesis by aortic rings in human blood: selective increase of prostacyclin mediated by a serum factor.

Synthesis of vascular epoprostenol (PGI2) and platelet thromboxane (TX) A2 is influenced by the coagulation cascade in incompletely understood ways. To elucidate this, prostanoids were determined in human blood anticoagulated by different drugs and incubated with and without rat aortic rings. Control incubations were performed in Hanks balanced salt solution. PGI2 and TXA2 synthesis were assessed by radioimmunoassay of their stable hydrolysis products 6-oxo-prostaglandin (PG) F1 alpha and TXB2. Fresh aortic rings incubated in Hanks solution with a thrombin inhibitor (TCK) synthesized similar quantities of 6-oxo-PGF1 alpha in the presence or absence of sodium citrate. In contrast, the intracellular calcium antagonist TMB-8 inhibited 6-oxo-PGF1 alpha synthesis. In contrast to the finding in Hanks solution, sodium citrate inhibited 6-oxo-PGF1 alpha synthesis by fresh aortic rings incubated in blood anticoagulated with TCK. However, TXB2 synthesis was not affected by citrate. Blood incubated alone at 37 degrees C in plain glass tubes generated a small amount of immunoreactive 6-oxo-PGF1 alpha. A thromboxane synthase inhibitor, OKY1581, increased immunoreactive 6-oxo-PGF1 alpha. However, blood anticoagulated with TCK and incubated similarly, generated no detectable 6-oxo-PGF1 alpha either in the presence or absence of OKY1581, showing that 6-oxo-PGF1 alpha synthesis in the previous experiments was dependent on the vascular rings. OKY1581 had little or no effect on 6-oxo-PGF1 alpha synthesis in incubations of fresh aortic rings with blood anticoagulated with TCK, despite inhibition of TXB2 synthesis. However, OKY1581 increased 6-oxo-PGF1 alpha synthesis by rings pretreated with acetylsalicylic acid (ASA) when incubated in blood, presumably by diversion of platelet endoperoxide to vascular PGI2 synthase. Sodium citrate did not influence the increase in 6-oxo-PGF1 alpha synthesis by ASA pretreated aortic rings caused by OKY1581 in whole blood. This implies that the PGI2 stimulating activity of whole blood in the absence of citrate exerts its effect proximal to PGI2 synthase. It is concluded that a low molecular weight serum factor formed during activation of the intrinsic coagulation pathway in blood, modulates PGI2/TXA2 balance by an action on vascular cyclo-oxygenase, possibly by an effect on intracellular calcium.