Evaluation of mammary blood flow measurements in lactating goats using the ultrasound Doppler principle

1. Non-invasive methods were developed for measuring mammary blood flow in lactating goats. 2. A Doppler principle ultrasound device was equipped with an external detector measuring maximal blood velocity (Vmax) and average blood velocity (Vav) was calculated as Vmax/2. Volume flow then depended on determination of the angle of insonation and the cross-sectional area of the milk vein (the caudal superficial epigastric or subcutaneous abdominal vein). 3. Blood velocities were measured on the milk vein of either side of the animal while clamping the pudendal veins manually. Blood velocities ranged from 7-34 cm/sec. 4. The milk vein diameter was measured by means of a slide gauge which, for clearly protruding veins, gave similar results to that measured by ultrasound scanning. In protruding veins the cross-section was circular. In non-protruding veins the cross-section was elliptical and the slide gauge significantly (P less than 0.01) overestimated the cross-sectional area. The milk vein diameter of either side measured in 10 lactating goats was 8.8 +/- 1.1 mm (means +/- SD). 5. Blood flow ranged from 90-675 ml/min in a dry and a high-yielding (3.4 l milk daily) goat, respectively. The reproducibility of the blood flow measurements was 12-16%. 6. It is concluded that the present method may be used for quantitative measurements of mammary blood flow in goats.     

Fluid volumes in rainbow trout, Salmo gairdneri: application of compartmental analysis

1. The measurement of fluid volumes by the indicator dilution technique and compartmental analysis was re-evaluated in free-swimming, undisturbed rainbow trout. 2. Plasma (33.5 ml/kg body wt) and blood (41.3 ml/kg body wt) volumes estimated by compartmental analysis from blood samples taken early (less than 5 min) after dye injection were 40% lower than volumes calculated by sampling late (greater than or equal to 80 min). 3. The rate of exchange of dye between plasma and interstitial fluid was high (48%/hr) compared to mammals (5%/hr) which supports the hypothesis that teleost capillaries have high protein permeability. 4. Total extracellular volume estimated using a single pool model (210.5 ml/kg body wt) of inulin kinetics was 20% higher than that calculated by a three pool model (172.8 ml/kg body wt).     

An improved simple technique for the collection of blood samples from rats and mice

The technique of blood collection from the lateral caudal vein has been improved. The method requires only moderate skill and no anaesthesia is necessary. Collection of blood samples causes little trauma and can be repeated at frequent (8 h) intervals thus making the method particularly suitable for pharmacokinetic and hormonal studies. Blood samples are uncontaminated by tissue fluids. Large volumes can be obtained, in mice up to 1.5 ml and in Wistar rats over 4 ml.     

Non-invasive blood sampling from primates using laboratory-bred blood-sucking bugs (Dipetalogaster maximus; Reduviidae, Heteroptera)

Primates are easily stressed by the conventional veterinary blood sampling routine and consequently, measured blood parameters may be biased. In this study, we tested blood-sucking bugs (Dipetalogaster maximus) on one lemur and two ape species (Microcebus murinus, Pongo abelii, Pan paniscus) as an alternative, non-invasive technique for bleeding primates. Within time periods of between 6 and 62 min we obtained blood volumes of 0.01–2.4 ml in 11 out of 12 trials from all three species. Therefore, we conclude that these bugs represent a new, gentle and effective tool for bleeding captive primates without stress.     

Comparison of Pathogen DNA Isolation Methods from Large Volumes of Whole Blood to Improve Molecular Diagnosis of Bloodstream Infections

For patients suffering from bloodstream infections (BSI) molecular diagnostics from whole blood holds promise to provide fast and adequate treatment. However, this approach is hampered by the need of large blood volumes. Three methods for pathogen DNA isolation from whole blood were compared, i.e. an enzymatic method (MolYsis, 1–5 ml), the novel non-enzymatic procedure (Polaris, 1–5 ml), and a method that does not entail removal of human DNA (Triton-Tris-EDTA EasyMAG, 200 µl). These methods were evaluated by processing blood spiked with 0–1000 CFU/ml of Staphylococcus aureus, Pseudomonas aeruginosa and Candida albicans. Downstream detection was performed with real-time PCR assays. Polaris and MolYsis processing followed by real-time PCRs enabled pathogen detection at clinically relevant concentrations of 1–10 CFU/ml blood. By increasing sample volumes, concurrent lower cycle threshold (Ct) values were obtained at clinically relevant pathogen concentrations, demonstrating the benefit of using larger blood volumes. A 100% detection rate at a concentration of 10 CFU/ml for all tested pathogens was obtained with the Polaris enrichment, whereas comparatively lower detection rates were measured for MolYsis (50–67%) and EasyMAG (58–79%). For the samples with a concentration of 1 CFU/ml Polaris resulted in most optimal detection rates of 70–75% (MolYsis 17–50% and TTE-EasyMAG 20–36%). The Polaris method was more reproducible, less labour intensive, and faster (45 minutes (including Qiagen DNA extraction) vs. 2 hours (MolYsis)). In conclusion, Polaris and MolYsis enrichment followed by DNA isolation and real-time PCR enables reliable and sensitive detection of bacteria and fungi from 5 ml blood. With Polaris results are available within 3 hours, showing potential for improved BSI diagnostics.     

Monitoring modulators of platelet aggregation in a microtiter plate assay

Platelets play a central role in maintaining biological hemostasis. Inappropriate platelet activation is responsible for thrombotic diseases such as myocardial infarction and stroke. Therefore, novel agents that can inhibit platelet activation are necessary. However, assays that monitor platelet aggregation are generally time-consuming and require high volumes of blood and specialized equipment. Therefore, a medium- to high-throughput assay that can monitor platelet aggregation would be considered useful. Such an assay should be sensitive, comparable to the "gold standard" assay of platelet aggregometry, and able to monitor multiple samples simultaneously but with low assay volumes. We have developed such a microtiter assay. It can assay an average of 60 independent treatments per 60 ml blood donation and demonstrates greater sensitivity than the current gold standard assay, namely platelet aggregation in stirring conditions in a platelet aggregometer. The microtiter plate (MTP) assay can detect known inhibitors of platelet function such as indomethacin, aspirin, and ReoPro. It is highly reproducible when using standard doses of agonists such as thrombin receptor-activating peptide (20 microM) and collagen (0.19 mg/ml). Finally, the MTP assay is rapid and sensitive and can detect unknown platelet-modulating agents from a library of compounds.     

Psychogenic and neurogenic effects on the intact forearm vein of man.

By means of an elegant venous preparation the behavior of intact veins of man can be studied decisively. The superficial veins of man are extremely sensitive to the environment and the autonomic and central nervous systems. This report illustrates the response of an isolated venous segment of the forearm to various psychogenic and neurogenic stimuli, with change in tone of the smooth muscle of the venous segment being reflected by change in segmental venous pressure. It is shown that the superficial forearm veins can be conditioned and that they are extremely dynamic and certainly not passive conduits for returning blood to the heart.     

Concentration of dilute solutions of macromolecules by electrophoresis

An electrophoresis apparatus which is used for concentrating micrograms of macromolecules from solutions as large as 250 ml is described. The recoveries were greater than 91% with three different macromolecules tested (28 to 360 kDa). Solutions with volumes in the range of 35 ml were concentrated 70-fold in less than 90 min to a final volume of 0.5 ml. Larger volumes in the range of 250 ml were concentrated 227-fold in 16 h to a final volume of 1.1 ml. Sterile concentrates can be obtained if the apparatus is constructed under sterile conditions.     

Bifurcation asymmetry of the porcine coronary vasculature and its implications on coronary flow heterogeneity

The branching pattern of the coronary arteries and veins is asymmetric, i.e., many small vessels branch off of a large trunk such that the two daughter vessels at a bifurcation are of unequal diameters and lengths. One important implication of the geometric vascular asymmetry is the dispersion of blood flow at a bifurcation, which leads to large spatial heterogeneity of myocardial blood flow. To document the asymmetric branching pattern of the coronary vessels, we computed an asymmetry ratio for the diameters and lengths of all vessels, defined as the ratio of the daughter diameters and lengths, respectively. Previous data from silicone elastomer cast of the entire coronary vasculature including arteries, arterioles, venules, and veins were analyzed. Data on smaller vessels were obtained from histological specimens by optical sectioning, whereas data on larger vessels were obtained from vascular casts. Asymmetry ratios for vascular areas, volumes, resistances, and flows of the various daughter vessels were computed from the asymmetry ratios of diameters and lengths for every order of mother vessel. The results show that the largest orders of arterial and venous vessels are most asymmetric and the degree of asymmetry decreases toward the smaller vessels. Furthermore, the diameter asymmetry at a bifurcation is significantly larger for the coronary veins (1.7-6.8 for sinus veins) than the corresponding arteries (1.5-5.8 for left anterior descending coronary artery) for orders 2-10, respectively. The reported diameter asymmetry at a bifurcation leads to significant heterogeneity of blood flow at a bifurcation. Hence, the present data quantify the dispersion of blood flow at a bifurcation and are essential for understanding flow heterogeneity in the coronary circulation.     

The influence of steroids on vascular tension of isolated superficial veins of the nose and face during the estrous cycle of gilts

The arrangement of the superficial facial veins enables blood flow from the nasal cavity into the peripheral circulation by two pathways: through the frontal vein into the cavernous sinus and through the facial vein into the external jugular vein. The current study was designed to determine whether estradiol and progesterone affect the vascular tone of the superficial veins of the nose and face in cycling gilts (Sus scrofa f. domestica) and to analyze the immunolocalization of progesterone receptors and estradiol receptors in these veins. The influence of hormones on vascular tension differed depending on the type of vessel and the phase of the estrous cycle. Estradiol decreased vascular tension in the nasal vein during the follicular phase (P < 0.05) and increased tension in the frontal vein during the luteal phase (P < 0.05). Progesterone increased the vascular tension of the frontal vein (P < 0.05) and decreased the tension of the other veins (P < 0.05) in both phases of the cycle. Expression of estradiol receptor β but not of progesterone receptor was observed in the superficial veins of the nose and face. In conclusion, the effect of ovarian steroid hormones on the vascular tension of the superficial veins of the nose and face in female pigs as well as the reactivity of these veins to steroid boar pheromones can affect the blood supply from the nasal cavity to the venous cavernous sinus. We propose that the ovarian steroid hormones that modulate the vascular tension of the nasal and facial veins may also influence the action of boar pheromones absorbed into the nasal mucosa in gilts and may reach the brain via local destination transfer.     

Method of high-precision microsample blood and plasma mass densitometry

The reliability of the mechanical oscillator technique (MOT) for blood and plasma mass density measurements on small samples is quantified in this paper. Sources of measurement errors that can reduce both the accuracy and precision of density determinations include storage of plasma samples, inhomogeneity of blood samples, and density reading before adequate temperature equilibration. Measurements on fractions from identical samples and repeated samplings from test subjects under steady-state conditions revealed a 10(-2) g/l reproducibility of density readings. The mean plasma density (PD) readings did not change significantly after up to 1-wk storage at +4 degrees C or up to 2 mo storage at -20 degrees C. The variability of the PD findings increased with storage time and were generally higher with storage at -20 degrees C, compared with +4 degrees C. Densitometers of different sizes were used to evaluate rheological influences on blood density (BD) readings. Linear correlations between PD and plasma protein concentration, between BD and blood hemoglobin concentration, and between erythrocyte density and mean corpuscular hemoglobin concentration were significant (P less than 0.001). Rapid density measurements with up to 10(-2) g/l reliability on small (less than 0.1 ml) volumes of biological fluids and continuous blood densitometry can be performed with use of the MOT.     

Tissue and plasma levels of endothelin in free flaps

The goal of the study was to assess whether endothelin-1 levels are increased in tissue and plasma in free flaps. To assess this hypothesis, blood samples were taken from the general circulation before and after reperfusion and from the flap after reperfusion in 20 patients undergoing breast reconstruction with free transverse rectus abdominis musculocutaneous or deep inferior epigastric perforator flaps. Tissue samples were also taken from the flap before and after the period of ischemia. Peripheral blood samples of 10 ml each were taken before the vessels were clamped and at 10 minutes and 1 hour after the flap was recharged. The flap vein was catheterized with a smooth catheter to avoid endothelial trauma, and ischemic blood from the flap was obtained immediately after the artery was unclamped and 10 minutes later. Two skin samples of 2 cm each were taken: one after dissection of the flap before division of the vessels and one after reanastomosis of the veins (one or two veins). Statistical analyses were performed with the (nonparametric) Wilcoxon signed rank test. Flap ischemia time, from vessel division to the completion of the arterial anastomosis, ranged from 35 to 120 minutes (mean, 48 minutes). The plasma endothelin-1 level extracted from the flap was 4.34 +/- 0.85 pg/ml, significantly higher than baseline, 3.87 +/- 0.81 pg/ml (p < 0.0001). There was a small increase, 4.5 +/- 1.03 pg/ml (p = NS), 10 minutes after reperfusion. The peripheral level after venous anastomosis was 3.78 +/- 0.79 pg/ml, not significantly different from the peripheral plasma level, before the flap was raised. The peripheral plasma level 1 hour after reperfusion was 3.83 +/- 0.8 pg/ml, with no difference from baseline. The tissue level of endothelin-1 before clamping was 3.8 +/- 0.8 pg/mg and in postischemic tissue, 5.2 +/- 0.6 pg/mg, a statistically significant increase. The authors concluded that endothelin-1 levels are elevated in free flaps. This could be an explanation for vasospasm and may lead to therapy directed against the no-reflow phenomenon.     

Cerebral blood flow assessment with indocyanine green bolus transit detection by near-infrared spectroscopy in the rat

In this study we evaluated the feasibility of measuring cerebral blood flow in rats by monitoring the transit of an indocyanine green bolus through the brain with multiwavelength near-infrared spectroscopy. Different volumes of a 1 mg/ml indocyanine green solution (5, 15, 25, 50 microl) were injected intravenously in the search for an optimal dose. Clear transit curves were obtained with all doses and a blood flow index could easily be determined. The indocyanine green signal obtained with the bolus of 5 microl rapidly returned to baseline and interfered minimally with the haemoglobin and cytochrome oxidase signals. This dose was used in a second study to evaluate the reproducibility of the signal and the effect of hypercapnia. Two groups of rats received 7 repetitive boli of indocyanine green. In one group, 7% CO(2) was added to the gas mixture before the second, fourth and sixth indocyanine green injection. Hypercapnia consistently caused a significant increase in blood flow index, cerebral haemoglobin concentration and O(2)-saturation. In the control group these variables remained stable in time. We conclude that monitoring of the transit of an indocyanine green bolus with multiwavelength near-infrared spectroscopy can be used to assess cerebral blood flow qualitatively in rats in combination with continuous monitoring of brain oxygenation.     

A microampere radiotelemetry transmitter for measurement of temperature from unrestrained subjects

A miniature radiotelemetry transmitter is described suitable for chronic or acute monitoring of temperatures from animal or human subjects. The transmitters can be easily constructed with little or no electronics background from discrete components readily available from any electronics supplier. Transmitters weighed from 2.5-20 g, had volumes of 0.8-3.5 ml, had normal currents of 50-120 muA with potential transmitter lives of 35 h to greater than 1 yr, and transmission ranges of 10-30 m. Transmitters could be implanted, swallowed, or connected to chronically or acutely attached temperature probes. The design of the telemetry system allows for reception on any commercial FM receiver. Depending on mode of decoding the temperature data transmitter signal can be accurate to measure changes of from 0.15 to 0.015 degrees C. Thus with a minimum of expense and little electronics background a temperature-dependent transmitter system can be constructed for monitoring temperatures from unrestrained subjects.     

小儿表浅静脉采集血标本成功率影响因素

目的：研究小儿表浅静脉采集血标本成功率影响因素,探讨提高表浅静脉采集血标本成功率的有效护理对策。方法：对每种影响因素进行分组研究,每组按随机分组原则设试验组和对照组,采取所需血量为成功,未采取所需血量为不成功。结果：常见几种影响因素试验组与对照组差异有统计学意义（p〈0.01）。结论：小儿表浅静脉采集血标本影响因素是多方面的,采血前必须对患儿进行综合评估,选择恰当的采血方法。     

Protein volumes and hydration effects. The calculations of partial specific volumes, neutron scattering matchpoints and 280-nm absorption coefficients for proteins and glycoproteins from amino acid sequences

Amino acid sequences, carbohydrate compositions and residue volumes are used to compare critically calculations of partial specific volumes v, neutron scattering matchpoints and 280-nm absorption coefficients with experimental v values for proteins and glycoproteins. The v values that are obtained from amino acid densitometry underestimate experimental v values by 0.01-0.02 ml/g while the v values from crystallographic volumes overestimate the experimental v values by 0.04-0.05 ml/g. An intermediate consensus volume set of amino-acid-residue volumes is proposed in order to predict experimental v values using sequence information. The method is extended to carbohydrates and glycoproteins. Neutron scattering matchpoints can be calculated from crystallographic residue volumes on the basis of the non-exchange of 10% of the main-chain NH protons. Crystallographic results on protein-bound water are used to account for the experimental values of v and matchpoints. Finally, 280-nm absorption coefficients, A1%, 1 cm 280, of 5-27 are found to be well predicted by the Wetlaufer procedure based on the totals of Trp, Tyr and Cys residues. Average errors are +/- 0.7, and the experimental A(1%,1cm)280 values can be larger than the predicted values by 3%.     

Detection of Bacteremia in Children with Sodium Polyanethol Sulfonate: a Prospective Clinical Study

A prospective study was made of 1,000 consecutive duplicate blood cultures obtained from sick children to evaluate the usefulness of sodium polyanethol sulfonate (SPS). With the small volumes of blood (1 to 5 ml) usually obtained for blood cultures in children, SPS did not increase the frequency of recovery of organisms judged to be associated with clinical infections, with the possible exception of Diplococcus pneumoniae. However, the use of SPS was associated with an increased frequency of recovery of organisms judged to be contaminants, such as Staphylococcus epidermidis and propionibacteria, possibly because SPS enhanced the recovery of a very small inoculum of skin bacteria.     

Physiological dead space during high-frequency ventilation in dogs

Tidal volumes used in high-frequency ventilation (HFV) may be smaller than anatomic dead space, but since gas exchange does take place, physiological dead space (VD) must be smaller than tidal volume (VT). We quantified changes in VD in three dogs at constant alveolar ventilation using the Bohr equation as VT was varied from 3 to 15 ml/kg and frequency (f) from 0.2 to 8 Hz, ranges that include normal as well as HFV. We found that VD was relatively constant at tidal volumes associated with normal ventilation (7-15 ml/kg) but fell sharply as VT was reduced further to tidal volumes associated with HFV (less than 7 ml/kg). The frequency required to maintain constant alveolar ventilation increased slowly as tidal volume was decreased from 15 to 7 ml/kg but rose sharply with attendant rapid increases in minute ventilation as tidal volumes were decreased to less than 7 ml/kg. At tidal volumes less than 7 ml/kg, the data deviated substantially from the conventional alveolar ventilation equation [f(VT - VD) = constant] but fit well a model derived previously for HFV. This model predicts that gas exchange with volumes smaller than dead space should vary approximately as the product of f and VT2.     

Quantification of 20alpha- and 20beta-dihydroprogesterone in plasma of the pregnant rhesus monkey

A competitive protein binding assay for 20α- and 20β-dihydroprogesterone is described which involves an initial chemical or enzymatic oxidation of these two isomers to progesterone. The assay can distinguish between 20α- and 20β-dihydroprogesterone and is sensitive to pg amounts of these two steroids. Venous steroid concentrations were measured in the pregnant rhesus monkey employing this assay. In this species the corpus luteum (CL) at days 22 and 157 of gestation is the primary ovarian source of 20α-dihydroprogesterone as indicated by a higher plasma concentration of this steroid in the ovarian vein draining the ovary containing the CL (+CL) than in the contralateral vein (?CL) (9.34 ng/ml versus 1.72 ng/ml, day 22; 7.52 ng/ml versus 1.96 ng/ml, day 157). By contrast the CL at day 50 appeared to secrete no 20α-dihydroprogesterone as evidenced by the essentially equal steroid levels in both ovarian veins. The CL synthesizes 20β-dihydroprogesterone only during early gestation (21–22 days) when the concentration of this steroid was 6.46 ng/ml and 0.87 ng/ml in the ovarian (+CL) and ovarian (?CL) veins, respectively. Synthesis of both 20α- and 20β-dihydroprogesterone occurs in the fetoplacental unit throughout pregnancy. This is indicated by higher steroid concentrations in the uterine vein than in the femoral vein. The results suggest both a qualitative and quantitative alteration in the luteal synthesis of 20α- and 20β-dihydroprogesterone with the advancement of gestation. The data also provide additional evidence that the steroidogenic activity of the CL is enhanced before parturition.     

Preparation of Oligonucleotides and Their Use in Molecular Weight Estimations

Abstract

Yeast RNA was used to prepare oligonucleotides employed to calibrate a G-50 Sephadex column. The oligonucleotides' preparation, isolation, desalting and characterization is described. Data obtained by chromatography of the oligonucleotides demonstrate that the molecular weights of oligonucleotides can be easily determined by interpolation using plots of elution volumes (Ve) versus molecular weights (M). Errors greater than 20% are obtained if the conventional plot of Ve-Vo/Vs versus log M is used (where Vo is the void volume of the column and Vs is the volume of the column occupied by the inert phase, the 6-50 Sephadex).