Whole Blood Measurements Research Articles

Reticulocytes and reticulated platelets: simultaneous measurement in whole blood by flow cytometry.

Reticulated platelets are a fraction of newly released circulating elements characterized by a residual amount of RNA. It has been suggested that the reticulated platelet count, providing an estimate of thrombopoiesis in the same way as erythrocyte reticulocyte count is a measure of erythropoiesis, may be useful in the study of thrombocytopenic disorders. Reticulated red cells and platelets can be analyzed by flow cytometry using specific stains for nucleic acids such as Thiazole Orange and Auramine-O. The aim of our work was to perform the simultaneous evaluation of reticulated elements in whole blood using a standard flow cytometer and to correlate the results obtained with a dedicated cytometer. A group of 14 patients with abnormal absolute reticulocyte counts (range 1.1-11%) and a group of 41 patients showing a platelet discrimination error when analyzed with a dedicated flow cytometer (Sysmex R1000) were enrolled. Linear amplification of both scatter and fluorescence was used to perform reticulocyte count. A gate was set on platelet dimensions, and logarithmic amplification of scatter and fluorescence was used to count reticulated platelets. A good correlation was obtained both for results of reticulocyte count (r2 = 0.9825) and for reticulated platelets (r2 = 0.8717) between our method and those using dedicated instruments. These data show that reticulated platelet count may be easily introduced in clinical laboratories that routinely perform reticulocyte count by flow cytometry.

CURRENT STATUS OF ACID-BASE QUANTITATION IN PHYSIOLOGY AND MEDICINE

Acid-base balance (ABB) is best determined by measurements in arterial whole blood, but for clinical purposes it can be expressed as estimated to apply to the entire extracellular fluid (ECF) of the body. ABB is determined by three independent variables: (1) respiratory (P co2); (2) metabolic (strong ion difference [SID]); (3) nonvolatile weak acid buffer (ATOT), primarily hemoglobin (Hb). Here we describe and compare four current indices of the metabolic component: (1) Standard base excess (SBE), in mM⋅L–1, quantifies the metabolic component, by assuming that the mean buffer strength of the ECF in humans equals that of blood with 5 g/dL hemoglobin. Despite individual variation of hemoglobin, the effective ATOT of ECF in vivo is practically invariant. SBE is automatically computed by most blood gas analyzers from P co2 and pH. (2) SID between plasma strong cations and anions, numerically equal to buffer base, as modified to apply to ECF in vivo. (3) HCO3– calculated from pH and P co2. Because pH and HCO3– vary with both P co2 and SID, equations are needed when this method is used to compare the sample with the usual or expected compensation for an ABB abnormality. Such computations do not produce a measure of the metabolic component. (4) Anion gap, Na+ + K+ − Cl– − HCO3–, detects anions not normally measured by plasma electrolyte screens. It can distinguish acidosis due to excess Cl– from that of other strong anions such as lactate. Using published pH (or H+) and HCO3– data, we have computed the typical compensatory responses of patients with primary acid-base imbalances in terms of SBE and Pa co2:

Blood compatibility and extended linearity of lactate enzyme electrode using poly(vinyl chloride) outer membranes

A functionally haemocompatible lactate enzyme electrode system with an extended linear range has been developed. The electrode is based on classical amperometric detection of hydrogen peroxide produced by the enzyme lactate oxidase, immobilised in a glutaraldehyde matrix. The sensor system comprises an outer poly(vinyl chloride), PVC, membrane with incorporated non-ionic surfactant which acts both as a haemocompatible interface and a diffusion barrier, extending the sensor linear range up to at least 90 mM lactate. Electroactive interferents, such as ascorbate, urate and acetaminophen, are effectively excluded by the further use of an inner sulphonated polyether-ether sulphone/polyether sulphone membrane. Membrane composition and fabrication were investigated. The blood compatibility of the outer PVC membranes was evaluated by comparing sensitivity of the electrode before and after exposure to undiluted whole blood. Scanning electron microscopy (SEM) was also used to examine the membrane surface. Measurements of whole blood and plasma lactate concentrations when compared with those obtained with a standard lactate analyser gave, respectively, y = 1.2x + 0.50, r 2 = 0.97, n = 6 and y = 0.89x + 0.13, r 2 = 0.96, n = 20.

Health Surveillance and Biological Effect Monitoring for Chromium-Exposed Workers

Although workers may be exposed to chromium metal, Cr(III) compounds, and Cr(VI) compounds at the workplace, only Cr(VI) compounds are of primary concern in terms of possible health hazards. A special health surveillance program must focus on known health impairments and target organs. Medical surveillance in combination with biological monitoring can help to protect the workers' health. Biological monitoring for chromium exposure in urine, blood, and erythrocytes provides different types of information. Whereas chromium measurement in urine and whole blood or plasma is indicative of recent total chromium exposure, chromium detection in erythrocytes is attributable to Cr(VI) only and covers retrospectively a longer period of time due to the erythrocyte life span. Possibilities of biological effect monitoring are discussed.

Heroin dependence effects on some major and trace elements.

In this study, 56 (14 control and 42 addicts) adult human subjects of both sexes of different periods of heroin dependence were subjected to the measurement of whole blood, serum, and red blood cell levels of some trace elements (zinc, manganese, iron copper, and bromine), as well as some major elements (phosphorus, sulfur, calcium, potassium, and chlorine). This was done by the energy-dispersive X-ray fluorescence (EDXRF) technique, in which copper and bromine showed a significant rise in whole blood (male) (22 and 32%, respectively), while zinc, iron, manganese, calcium, sulfur phosphorus, potassium, and chlorine showed a significant drop (49, 8, 25, 34, 21, 51, 61, and 72%, respectively) in proportion to the period of heroin intake (6 yr) and in comparison with the control group. No significant sexual variation has been reported.

Stabilized needle electrode system for in vivo glucose monitoring based on open flow microperfusion.

Preliminary in vitro studies and in vivo performance of amperometric glucose needle enzyme electrodes incorporating an open microflow technique, in which the sensor surface in subjected to a flow of fluid, are reported. Initially using a slow flow (60 microliters h-1) of isotonic phosphate buffer over the enzyme electrode tip, an interface was created which reduced cellular/protein fouling for electrode measurements in whole blood. Here a minor reduction in electrode response (apparent only at high glucose concentration) occurred which was not cumulative and therefore not associated with fouling. The protection afforded by the moving aqueous film was independent of fluid composition; the use of isotonic/hypertonic buffer, addition of anticoagulant (1% m/v heparin) or enhanced fluid viscosity (addition of 1% v/v glycerol) did not affect the system. Implantation of the electrode and its microflow cannula into subcutaneous tissue in rats was associated with a decreased buffer flow (30 microliters h(-1)) and had the effect of (i) reducing electrode stabilization (30 min), (ii) accelerating 'pick up' of tissue glucose changes after intravenous glucose (1-2 min lag) or insulin (3-7 min lag) and (iii) achieving a correlation between tissue and blood glucose values under dynamic conditions (r2 = 0.98, y = 0.99x + 0.23). Reassessment of the electrode response in vitro, following a 4 h monitoring period, provided a sensor response within 3% of the original electrode sensitivity, indicating little or no surface fouling and avoiding the requirement for repeated in vivo calibrations at least over the initial implantation period.

More on the measurement of ionized magnesium in whole blood.

Current technology has made it possible to measure ionized magnesium with user-friendly ion selective electrode technology. Although this technology has not reached a perfect status literature shows that it can be used in clinical routine especially in serum. This paper deals with issues concerning the measurement of ionized magnesium in whole blood. To measure in whole blood apart from an instrument with good general performance data an adequate sample treatment is especially critical. We tested a number of different sample containers and found that they interfere in different degrees with the magnesium determination. In some cases this is due to silicone giving falsely high ionized magnesium values which may be twice the real value and even higher. Another problem is the heparinization of the sample. We found that different heparins may alter the result in a different way. The well known complexing capability of heparin may lead to a reduction of the ionized magnesium within the sample and thus to false low results. But some heparins cause elevated results. The reason for this may be zinc. Test procedures to check the quality of anticoagulants are given. Due to these procedures sample containers could be found which do not interfere with the determination of ionized magnesium in whole blood - thus making the measurement possible. All these factors indicate that the measurement of ionized magnesium really needs a very well defined sample handling - otherwise false results may arise. This is not only true for the measurement of whole blood but also for plasma or even for "so called serum" as there might be a sample container within the chain from patient to the instrument which is unsuitable- and thus will alter the result.

Spectral biodiagnostics of tissues with fiber optics

AbstractThe synergy of novel MIR‐fibers with FTIR‐spectroscopy opens a new horizon in applications of spectroscopy in 4 − 16 μm range in vivo medical diagnostics. The innovated bare core and core‐clad MIR‐fibers are produced by hot extrusion method from the crystals of silver halides solid solutions. These non‐toxic and non‐hygroscopic fibers are characterized by low optical losses: 0.1 − 0.5 dB/m nearby 10μm, and high flexibility: Rbending > 10–100 fiber diameters. Some prototypes of accessories for FTIR‐spectrometers, based on commercially available MIR‐fibers for several areas of promising applications are developed. Non‐invasive medical diagnostics in vivo: cancer detection and glucose content measurements in whole human blood is possible. The new family of evanescent fiber sensors is started for the direct in‐process applications. Fiberoptic Evanescent Wave FTIR Spectroscopy (FEWS) for cancer diagnosis in human organs is exemplified in this paper, and their great potentiality is demonstrated.

Methylmercury: significance of intrauterine and postnatal exposures

Outbreaks of methylmercury poisoning in Japan and Iraq have demonstrated the sensitivity of the fetus to neurotoxic effects. Based on toxicokinetics and considerations of practicability, the optimal biomarker of methylmercury exposure is the hair concentration, but whole-blood measurements of mercury are also useful. Dose-response relations are still incompletely known, especially concentrating developmental neurotoxicity under conditions of chronic exposure. Available evidence indicates that neurobehavioral dysfunction in children may occur if the maternal mercury concentration in hair is > 6 micrograms/g (30 nmol/g). This value corresponds to a blood mercury concentration of approximately 24 micrograms/L (120 nmol/L). The period of maximum sensitivity of the nervous system to methylmercury toxicity is unknown, but the transfer of mercury to the newborn through human milk may represent an additional risk. In view of the wide occurrence of mercury contamination in developing countries, increased use of the exposure biomarkers is encouraged.

Amperometric Glucose Biosensor for an Undiluted Whole-Blood Analysis

The design of an enzymatic biosensor based on the use of immobilized glucose oxidase is described for the determination of glucose in blood serum and undiluted whole blood. Measurements in whole blood with RSD=0.3% are possible due to the use of an outer polypropylene membrane wetted with a mixture of various polyoxyethylene ethers (Triton X-100).

Some Pitfalls in the Measurement of Blood Glutathione

1. We report some pitfalls in the measurement of whole blood and plasma glutathione in man. 2. Using a simple tourniquet to the forearm and a 21-gauge needle, blood samples were collected by brachial vein puncture from healthy subjects. Whole blood and plasma were analysed for total glutathione, including the reduced and oxidized forms, by a spectrophotometric recycling method involving the glutathione reductase/NADPH couple. 3. The concentration of oxidized glutathione was determined after treatment of aliquots with either 2-vinylpyridine or N-ethylmaleimide to trap reduced glutathione. Reduced glutathione in the native samples could then be obtained by subtraction. 4. When the reagents were added to separated plasma, 2-vinylpyridine yielded values for oxidized glutathione that were twice as high as with N-ethylmaleimide. In whole blood studies the discrepancy was even greater, and the problem was not resolved by deproteinization of samples with 5-sulphosalicylic acid. Using N-ethylmaleimide, levels of oxidized glutathione were less than 1% of total glutathione in whole blood. 5. Despite attempts to minimize haemolysis, lysed erythrocytes contributed on average 25% to the 'plasma' glutathione concentration.

Comparison of Cyclosporine Measurement in Whole Blood by High-Performance Liquid Chromatography, Monoclonal Fluorescence Polarization Immunoassay, and Monoclonal Enzyme-Multiplied Immunoassay

Monitoring of cyclosporine concentrations in whole blood is used routinely as a guide to adjusting dose so as to achieve optimal therapeutic benefit with minimal adverse effects. In the present study, we have compared a specific high-performance liquid chromatography (HPLC) assay with a fluorescence polarization immunoassay (TDx) and an enzyme-multiplied immunoassay (Emit). Both Emit and TDx assays employ a monoclonal antibody to cyclosporin A and therefore have the potential for a high degree of specificity. Blood specimens (EDTA as anticoagulant) were obtained from 113 patients (71 renal transplants, 17 liver transplants, and 25 other categories) taking cyclosporine and analysed by all three methods. There were significant correlations between results for HPLC and Emit (Emit = 10.54 + 1.07 x HPLC; r2 = 0.82, p less than 0.001) and between results for HPLC and TDx (TDx = 9.16 + 1.42 x HPLC; r2 = 0.82, p less than 0.001). Compared to HPLC analysis, 74% and 96%, respectively, of Emit and TDx results were to the left of the line of identity. The TDx monoclonal antibody appears to have a lesser degree of specificity than that used in the Emit assay. Mean concentrations of cyclosporine measured by Emit and TDx were 17% and 51% higher, respectively, than those measured by HPLC. Because of this overestimation, we suggest that both Emit and TDx methods may find their most appropriate use in routine therapeutic monitoring of renal transplant patients in whom metabolite concentrations are less variable over time.

Iron-dextran as a magnetic susceptibility contrast agent: flow-related contrast effects in the T2-weighted spin-echo MRI of normal rat and cat brain.

Iron-dextran (1 mmol Fe/kg) was used as an intravascular, paramagnetic contrast agent in rat and cat brain in conventional spin-echo T2-weighted (TR 2800/TE 100) 1H magnetic resonance imaging. The resulting images displayed differential decreases (30-50%) in intensity whose pattern was similar to that obtained with the superparamagnetic particulate iron oxide AMI-25 (0.18 mmol Fe/kg). Postcontrast images displayed improved anatomic detail, and contrast effects were observed to be greater in cortical and subcortical gray matter than in adjacent white matter. Intravenous injection of acetazolamide after administration of iron-dextran caused a small additional decrease in image intensity. Measurement of whole blood and plasma at 5 min postinjection of either contrast agent revealed significant increases in their volume magnetic susceptibilities. The contrast effect appears to be related to magnetic susceptibility changes brought about by the iron-dextran; it has both blood volume and blood flow components. The static model of magnetic susceptibility effects in brain capillaries is modified to include bolus flow of erythrocytes, providing a mechanism for the observed flow effects.

Rheological measurements of whole blood from Alzheimer's disease, dialysis and multinfarctual dementia patients: a,eP.E. Zatta, bA.M. Tollardo, cM. Favarato, dG. Bazzato, dP. Moracchiello d, b,eM.Nicolini. aCentroCNR,Emocianine,Metalloproteine, bCUGAS Universita' di Padova, cDipartimento di Biologia, dDivisione di Nefrologia, Ospedale Civile di Mestre-Venezia, eDipartimento di Scienze Farmaceutiche, Italy

Rheological measurements of whole blood from Alzheimer's disease, dialysis and multinfarctual dementia patients: a,eP.E. Zatta, bA.M. Tollardo, cM. Favarato, dG. Bazzato, dP. Moracchiello d, b,eM.Nicolini. aCentroCNR,Emocianine,Metalloproteine, bCUGAS Universita' di Padova, cDipartimento di Biologia, dDivisione di Nefrologia, Ospedale Civile di Mestre-Venezia, eDipartimento di Scienze Farmaceutiche, Italy

Histamine measurement in whole blood and cell supernatants

Histamine as an important mediator of the allergic reaction has been measured by a number of methods both in whole blood and cell supernatants after in vitro release. We compared the fluorometric histamine assay with two radioimmunoassays and studied detection limit, recovery, cross-reactivity and accuracy in buffer-based standards as well as the sensitivity after IgE-mediated in vitro release from washed cells and whole blood. Our data indicate that the double-antibody RIA (Pharmacia) can be used for studies of basophil histamine release in both whole blood and cell supernatants. Due to significant cross-reactivity to N-methyl-histamine it cannot be used for histamine determinations in plasma. The second radioimmunoassay using succinyl-glycine as an acylating reagent and monoclonal antibodies, which have been raised to acylated histamine, is the most sensitive assay without any cross-reactivity with methylhistamine.

Performance Characteristics of Three Whole-Blood Glucose Monitors

We evaluated the performance of three whole-blood glucose meters for introduction to hospital bedside testing. The samples were from hospital patients who had a wide range of glucose concentrations and hematocrit values. The whole-blood specimens were kept on ice to minimize preanalytic degradation. Plasma glucose from each sample was quantitated for the reference value. The maximum acceptable error between whole-blood measurements and reference was set at ±15%. Glucose meter readings exceeded this limit in 35.4%, 27.0%, and 43.4% of samples with the three different meters. Variation from reference values was also analyzed according to specimen hematocrit and glucose concentration. Each instrument exhibited different bias with hematocrit and glucose. These results indicate that none of the three instruments has met current standards for interchangeable hospital-wide use.

Thirty-Second Sampling of Plasma Growth Hormone in Man: Correlation with Sleep Stages*

Both fasting and sleep increase the secretion of human GH and, therefore, might explain its predominantly nocturnal release. To study the precise temporal relationship between GH secretory episodes and cortical activity, GH measurements and electroencephalogram sleep stage recordings were performed every 30 s for 8 h in six young male volunteers fasted for 24 h. GH was measured in two drops of whole blood, which were directly sampled into the assay tube using a continuous blood withdrawal pump and a fraction collector. Concomitant serum sampling during a GH-releasing hormone test (n = 4) revealed a high correlation (r = 0.98) between GH measurements in serum and whole blood. GH pulses were objectively identified with Cluster analysis, and GH secretion rates were calculated with a waveform-independent deconvolution algorithm. When data were analyzed as replicates with 1-min intervals, the nocturnal pulse frequency was 1.2 pulses/h. Elimination of data points demonstrated 43% and 64% reductions in the number of GH pulses detected for 5- and 20-min sampling intervals, respectively. Mean GH concentrations and secretory rates were significantly higher during stage 3 and 4 sleep compared to stage 1, 2, and rapid eye movement sleep. GH secretory rates and peripheral GH concentrations were maximally correlated with sleep stage, with lags of 4.5 and 16 min, respectively, suggesting that maximal GH release occurs within minutes of the onset of stage 3 or 4 sleep. This temporal coincidence between pituitary GH secretion and delta sleep is consistent with cortical control over hypothalamic-pituitary function.

Fluorescence Polarization Immunoassay for Cyclosporine A Determination in Whole Blood

A fluorescence polarization immunoassay (FPIA) for cyclosporine A (CsA) determination in blood was evaluated. The method demonstrated a good reproducibility both in within-run and between-run assays. The linearity range was 35-990 micrograms/L. Comparison of measurements in patients' whole blood with radioimmunoassay results showed higher estimates by the FPIA despite the good correlation coefficient. Finally, a poor agreement was observed between the evaluated method and the high-performance liquid chromatographic technique for CsA determination.

Carcinoid syndrome and serotonin: Therapeutic effects of ketanserin

The carcinoid syndrome can arise when effluent blood from carcinoid tumor tissue gains access to the systemic, as opposed to the portal, venous system. Features include facial flushing, diarrhea, wheezing, right-sided cardiac lesions, and retroperitoneal fibrosis. Attacks of flushing, diarrhea, and wheezing can be provoked by bolus injections of adrenaline, noradrenaline, or pentagastrin. While serotonin usually predominates, carcinoid tumors can also secrete, in varying proportions, 5-hydroxytryptophan, kallikrein, kinins, substance P and other neuropeptides, prostaglandins, catecholamines, and histamine. Of these, serotonin, kinins, histamine, and substance P are possible mediators of flushes; serotonin and substance P of hyperperistalsis; and serotonin, kinins, or histamine of bronchial constriction. Despite the gross excess of circulating serotonin, nearly all is platelet bound and therefore inactive. Very little is free in plasma. Demonstration of a contribution of serotonin to carcinoid attacks requires assay of free plasma serotonin; measurements of whole blood or serum serotonin are of little value. Some, but not all, provoked flushes have been shown to be accompanied by a rise in free plasma serotonin or substance P; an increase in circulating kinins has been more consistently shown. The 5HT2 antagonist ketanserin has been found to inhibit both provoked and spontaneous attacks of flushing, diarrhea, and dyspnea in a proportion of patients with carcinoid syndrome.(ABSTRACT TRUNCATED AT 250 WORDS)

Measurement of 5-Methyl tetrahydrofolate (5-Methyl THF) by high performance liquid chromatography (HPLC)

Measurement of serum and red cell folate is one of the commonest tests in Haematology. The methods in current use are microbiological (L.casei) and radioisotope dilution assay (RDA). Microbiological assays are slow and subject to interference from antibiotics and other drugs. RDA are expensive and require isotope counting equipment. In addition, although the assays have been clinically useful, quality assurance trials have shown poor precision and poor agreement between different laboratories. Neither assay is specific for 5-Methyl THF in serum by HPLC and that measurement of whole blood and red cell 5-Methyl THF is even easier by this method. Our preliminary results show precision comparable with existing methods and correlate well with these methods. Another benefit is that it is possible to measure other folate species, which may provide further insight into folate metabolism.