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PubMed1998

Comparative recoveries of Naegleria fowleri amoebae from seeded river water by filtration and centrifugation.

Detection of pathogenic Naegleria fowleri in environmental water samples, which is necessary for the prevention of primary amoebic meningoencephalitis, generally requires concentrating the samples. Two concentration techniques, filtration and centrifugation, were used to study the recovery of N. fowleri, in vegetative or cystic form, that had been mixed with the two other thermotolerant Naegleria species, N. lovaniensis and N. australiensis. Counting of amoebae was performed by the most probable number method on 10 water replicates of 100 ml and 10 ml each. With both concentration methods, recovery was better for cysts than for trophozoites (53% +/- 21% versus 5% +/- 5% by filtration and 57% +/- 25% versus 22% +/- 5% by centrifugation). The recovery of Naegleria trophozoites by filtration was very low, and centrifugation was significantly better than filtration in recovery of Naegleria trophozoites (22% +/- 5% versus 5% +/- 5%; P < 0.001). For cysts, however, filtration appeared as efficient as centrifugation, with equivalent values for recovery (53% +/- 21% versus 57% +/- 25%; P > 0.7). Although the recovery of cysts of N. fowleri obtained by filtration (51% +/- 24%) appeared higher than that by centrifugation (36% +/- 23%), the difference was not significant (P > 0.1). Both concentration methods have highly variable recovery rates, making accurate quantification of low concentrations (< 100/liter) of N. fowleri in the environment difficult.

Animals↗
PubMed1988

Effect of dexamethasone on detection of herpes simplex virus in clinical specimens by conventional cell culture and rapid 24-well plate centrifugation.

During a 4-month period, two methods for rapid detection of herpes simplex virus (HSV) were examined: (i) pretreatment of A549 cells with dexamethasone for conventional tissue culture (277 specimens) and (ii) 24-well plate centrifugation using A549 cells with and without dexamethasone pretreatment and staining with serotype-specific monoclonal antibodies (Syva Co., Palo Alto, Calif.) after incubation for 16 to 18 h (153 specimens). By conventional tube cell culture, both with and without dexamethasone, HSV was identified in 88 of 277 (32%) specimens. Significantly more specimens were positive for HSV at 24 h (46 versus 27 specimens) and at 48 h (a total of 72 versus 59 specimens) (P less than 0.0001) in dexamethasone-treated A549 cells. Of the 153 specimens tested by conventional culture and 24-well plate centrifugation, HSV was detected in 44 (29%) by conventional culture, and by 24-well plate centrifugation with and without dexamethasone, HSV was detected in 32 (21%) and 30 (20%) specimens, respectively. The sensitivity, specificity, and positive and negative predictive values of 24-well plate centrifugation with A549 cells for detection of HSV were 73 (71% without dexamethasone), 100, 100, and 90%, respectively. In conventional tube cell culture, pretreatment of A549 cells with dexamethasone results in more rapid detection of HSV. Centrifugal inoculation of dexamethasone-treated and untreated A549 cells in 24-well plates and staining with monoclonal antibodies after incubation for 16 to 18 h is an insensitive means to detect HSV in clinical specimens and should not replace conventional tube cell culture.

Cells, Cultured↗
PubMed1979

Improved blood culture technique based on centrifugation: clinical evaluation.

A total of 3,335 blood samples from 1,180 patients suspected of having bacteremia were analyzed concurrently by two methods: (i) supplemented peptone broth with sodium polyanethanol sulfonate and a CO2 atmosphere; and (ii) lysis centrifugation at 3,000 X g for 30 min onto a high-density, hydrophobic cushion. The centrifugation technique recovered 80% of the positive cultures as compared with 67% for the broth method. The centrifugation technique showed an apparent increase in the isolation of staphylococcus aureus, Pseudomonas, and yeasts. In almost every instance, the time required for detection of a positive culture was shortest for the centrifugation method. Contamination rates for both systems were comparable (1.4%). Quantitation, offered only by the centrifugation method, proved useful on several occasions in discriminating between an opportunistic infection versus a skin contaminant and in judging efficacy of antimicrobial therapy.

Bacteria↗
PubMed1968

Density gradient centrifugation of rubella virus.

Rubella virus was centrifuged in sucrose density gradients. One of two densities could be ascribed to the virus, depending upon the suspending medium used. The virus was found at a density of 1.16 g/cm(3) after centrifugation for 18 hr in sucrose gradients prepared in distilled water. By contrast, when the sucrose gradients were prepared in tris(hydroxymethyl)aminomethane (Tris)buffer containing ethylenediaminetetraacetic acid (EDTA), the virus was found at a density of 1.18 g/cm(3) after 18 hr of centrifugation. The virus banded at this higher density after only 2 hr of centrifugation when pretreated by overnight incubation in the Tris-EDTA buffer. A kinetic study showed that, in sucrose gradients containing this buffer, the virus gradually migrated as a single peak of infectivity from a density of 1.16 g/cm(3) after 2 hr of centrifugation to the higher 1.18 g/cm(3) density after 18 hr. The density change was shown to be reversible; after the removal of the Tris-EDTA buffer, rebanding of virus harvested at the heavy density resulted in its banding at the lower 1.16 g/cm(3) density. The data indicate that density change could not be explained on the basis of the loss of some component from the virus or on the basis of the failure of the virus to reach equilibrium. However, it is possible that the two densities observed were a reflection of the existence of rubella virus in different hydration states in the presence and absence of Tris buffer containing EDTA.

Centrifugation, Density Gradient↗
PubMed1968

Enhancement of infectivity of murine cytomegalovirus in vitro by centrifugal inoculation.

Centrifugation of murine cytomegalovirus inocula from a variety of sources onto secondary mouse embryo cell monolayers at 1,900 x g for 30 min regularly revealed 10- to 100-fold more infectious virus than could be found in the same materials using standard inoculation methods. Virus demonstrable only by centrifugation was present throughout the entire growth cycle in a constant proportion to virus measured without centrifugation. Extracellular growth curves of both populations revealed an 18- to 21-hr latent period, followed by a long-linear increase over the next 12 hr; final yield was 30 plaque-forming units (PFU) per cell. Centrifugation of cells prior to inoculation or after standard adsorption and removal of inoculum failed to result in any significant change in measured virus titer. However, even after 4-hr adsorption, the supernatant inoculum could be transferred and centrifuged onto a fresh monolayer resulting in the same increment of measurable virus. Neutralizing antibody and interferon were equally efficacious against 100 PFU of virus as defined by either method. Thus, this newly identified population of cytomegalo-virus represents the vast majority of potentially infectious units and appears to differ solely in ease of adsorption onto cell monolayers.

Adsorption↗
PubMed1990

Stimulation of proteoglycan and DNA syntheses in chondrocytes by centrifugation.

So that the effects of biomechanical forces on the proliferation of chondrocytes and their proteoglycan synthesis could be studied, growth-plate and articular chondrocytes were maintained separately as packed masses in centrifuge tubes in the presence of 10% serum. In these conditions, the cells became re-organized into cartilaginous tissue in seven days. After ten days, they were centrifuged at gravities (g) of 1.3-27 for 24 h in a CO2 incubator. Control cells were maintained in the CO2 incubator without centrifugation. Centrifugation of growth-plate chondrocytes at 3 g resulted in a two-fold increase in incorporation of [35S]sulfate into proteoglycans, but had little effect on their [3H]thymidine incorporation into DNA. On the other hand, centrifugation of articular chondrocytes at 3 g for 24 h caused 1.5-fold increases in both [35S]sulfate incorporation into proteoglycans and [3H]thymidine incorporation into DNA. These results suggest that biomechanical forces have different effects on the growth and differentiation of articular and growth-plate chondrocytes.

Animals↗
PubMed1992

High-flow femoro-femoral bypass utilizing small cannulae and a centrifugal pump on the venous side.

Femoro-femoral bypass is an established technique in the armamentarium of cardiac surgeons, but poor venous drainage usually restricts the flow rate that can be achieved. We describe a technique whereby full flow ( greater than 2.41 l/min/m 2 femoro-femoral bypass) can be achieved with a 17 F arterial and a single 21 F venous cannula placed percutaneously or via a cut-down. Transoesophageal echo is used to position the tip of the venous cannula accurately in the right atrium. The circuit includes a centrifugal pump on the venous side, pumping into a reservoir; a conventional roller pump delivers blood through the arterial cannula. A parallel arrangement allows the centrifugal pump to be excluded from the circuit at any stage. The system allows flow rates over 2.4 l/min/m 2 despite the size of the venous cannula; without the centrifugal pump working maximal flow rates are under 1.5 l/min/m 2. The right side of the heart is totally decompressed and there is no need to add volume or vasopressors to maintain the desired full flow rate. Once the chest is open, perfusion may continue as before or gravity drainage can be utilized after stopping the centrifugal pump; venous return may be augmented by placing additional cannulae. If desired, slowing drainage by the centrifugal pump temporarily permits the blind placement of a coronary sinus cannula without entraining air.(ABSTRACT TRUNCATED AT 250 WORDS)

Blood Pressure↗
PubMed1991

Filtering centrifugation through two layers of silicone oil: a method for the kinetic analysis of rapid metabolite transport in organelles.

Kinetic studies of ATP uptake in amyloplasts from sycamore (Acer pseudoplatanus L.) have been performed with a newly developed method of centrifugation through a double layer of silicone oil; the results are compared with the frequently used method of centrifugation through a single layer. The present technique employs two separate silicone layers: the upper one prevents mixing of the organelle preparation with the incubation layer, which contains the metabolites, and the lower one prevents mixing of the incubation and pelleting layers. Incubation of the organelles takes place in the incubation layer after the upper layer of silicone inverts during centrifugation. Depending on the speed of centrifugation, incubation periods as short as 1 sec and as long as 2 min can be achieved. High reproducibility and accuracy, acquisition of multiple data in a single centrifugation, and maintenance of structural integrity and metabolic activity of the organelles make the technique advantageous for the analysis of metabolite transport in amyloplasts and other kinds of organelles.

Adenosine Triphosphate↗
PubMed1988

Development of centrifuged cow zygotes cultured in rabbit oviducts.

Zygotes from superovulated cows were centrifuged and pronuclei were detected by differential interference-contrast microscopy in 73% of 106 zygotes. Zygotes were then transferred to ligated oviducts of follicular-phase, 1-day pseudopregnant or 7-day pseudopregnant rabbits and recovered 5 days later. Their development did not differ from that of uncentrifuged zygotes transferred to the opposite oviduct: 41% of the embryos recovered from rabbit oviducts contained 17-32 nuclei and an additional 5% contained greater than 32 nuclei. In another experiment, 399 ova from unmated cows were transferred to rabbit oviducts to determine whether centrifugation induced parthenogenetic development. After 7 days, 257 ova were recovered; 16% of the recovered ova had developed parthenogenetically and contained 2-30 nuclei. Neither centrifugation of the ova nor reproductive status of the rabbits influenced the proportion of parthenogenotes found. Parthenogenetic development was also observed in 14 of 71 ova (20%) recovered on Day 7 from uninseminated superovulated cows. In an attempt to increase the probability of detecting treatment differences, centrifuged and control cow zygotes were incubated for 7 (rather than 5) days in opposite oviducts of fourteen 1-day pseudopregnant rabbits. Development was unaffected by centrifugation: 61% of the zygotes recovered had developed beyond the 16-cell stage, with 23, 24 and 15% containing 17-32, 33-64, and greater than 64 nuclei, respectively. Taking into account the percentage of zygotes in which pronuclei can be seen, the recovery rate from rabbit oviducts, and the proportion of embryos that develop to the morula stage or beyond, 26% of the original group of zygotes would be candidates for transfer into recipient cows.

Animals↗
PubMed1990

[Evaluation of separating X- and Y-sperms by percoll density gradient centrifugation using sperms of transgenic mice carrying a transgene on Y-chromosome].

Using sperms of the transgenic mice carrying a human A gamma/beta-globin gene on Y-chromosome, we attempted to separate X- and Y-bearing sperms by the Percoll density gradient centrifugation. The ratio of X- and Y-sperms was determined by DNA dot blot hybridization procedure with sperm DNA. Sperm suspension collected from cauda epididymidis was loaded on the gradient composed of 7 Percoll concentrations (35-84%) and was centrifuged at 300 x g for 10, 15 or 20 minutes, respectively, at room temperature. After centrifugation, sperms were collected from each gradient fraction and washed with 0.85% saline solution. DNA was extracted from sperms, dotted and fixed on nitrocellulose filter, and was hybridized with the 32P-labeled DNA probe derived from the beta-globin gene. Each DNA spot was cut out, immersed in the liquid scintillator and was counted for radioactivity. There was no difference among the radioactivities in the DNA spots, indicating that the ratio of X- and Y-sperms was the same in all the gradient fractions of three different centrifugal conditions. The results suggests to be difficult to separate X- and Y-sperms by Percoll density gradient centrifugation, at least, using sperms from cauda epididymidis of mouse.

Animals↗
PubMed2000

Evaluation of an in-house centrifugal hematology analyzer for use in veterinary practice.

OBJECTIVE: To compare CBC results obtained by use of an in-house centrifugal analyzer with results of a reference method. DESIGN: Prospective study. SAMPLE POPULATION: Blood samples from 147 dogs, 42 cats, and 60 horses admitted to a veterinary teaching hospital and from 24 cows in a commercial dairy herd. PROCEDURE: Results obtained with the centrifugal analyzer were compared with results obtained with an electrical-impedance light-scatter hematology analyzer and manual differential cell counting (reference method). RESULTS: The centrifugal analyzer yielded error messages for 50 of 273 (18%) samples. Error messages were most common for samples with values outside established reference ranges. Correlation coefficients ranged from 0.80 to 0.99 for Hct, 0.55 to 0.90 for platelet count, 0.76 to 0.95 for total WBC count, and 0.63 (cattle) to 0.82 (cats) to 0.95 (dogs and horses) for granulocyte count. Coefficients for mononuclear cell (combined lymphocyte and monocyte) counts were 0.56, 0.65, 0.68, and 0.92 for cats, horses, dogs, and cattle, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that there was an excellent correlation between results of the centrifugal analyzer and results of the reference method only for Hct in feline, canine, and equine samples; WBC count in canine and equine samples; granulocyte count in canine and equine samples; and reticulocyte count in canine samples. However, an inability to identify abnormal cells, the high percentage of error messages, particularly for samples with abnormal WBC counts, and the wide confidence intervals precluded reliance on differential cell counts obtained with the centrifugal analyzer.

Animals↗
PubMedDate not supplied

Combination of direct swim-up technique and discontinuous Percoll gradient centrifugation for sperm preparation of oligoasthenozoospermic samples.

Sperm recovery for assisted reproduction in oligoasthenozoospermic patients is not satisfying either by the swim-up technique or by Percoll gradient centrifugation, and no single technique is constantly preferred. The design of this study was to evaluate the effects of combining the two methods on improving the efficacy of sperm preparation in these poor samples. For each semen sample, 1 mL was treated with a combination method, which used direct swim-up technique to recover motile sperm swimming to the supernatant, and then the residual semen was subjected to two-layer discontinuous Percoll gradient procedure for further recovery of motile sperm. Another 1 mL was prepared with two-layer discontinuous Percoll gradient centrifugation alone for comparison. Parameters measured included sperm concentration, number of progressively motile sperm, percentage of progressive motility, percentage of motile sperm recovery, amount of debris, percentage of normal forms according to Kruger's strict criteria, and motion characteristics of sperm using computer-aided motility analysis. The results of 30 oligoasthenozoospermic samples demonstrated that the combination method achieved a significantly greater recovery of motile sperm than the two-layer discontinuous Percoll gradient centrifugation did (43.2 +/- 19.7% vs. 32.2 +/- 14.3%, p < .05). The percentage of progressive motility was higher in the samples of the combination method than in those of Percoll gradient centrifugation alone, but the difference was not significant (63.7 +/- 21.8% vs. 58.7 +/- 20.1%). The debris of semen was removed equally well by both methods. The percentage of normal forms as well as motion characteristics, including curvilinear velocity, straight-line velocity, mean amplitude of lateral head displacement, and linearity, were similar in the samples treated by these two procedures. The combination of the direct swim-up technique and discontinuous Percoll gradient centrifugation combines the advantages of each procedure and results in greater recovery of motile sperm in oligoasthenozoospermic samples.

Cell Separation↗
PubMed1989

Effects of centrifugation on transmembrane water loss from normal and pathologic erythrocytes.

Plasma 125I-albumin was used as a marker of extracellular dilution in order to study the effect of high-speed centrifugation on transmembrane water distribution in several types of human red cells, including normal (AA), hemoglobin variants (beta A, AS, SC, beta S, and SS), and those from patients with hereditary spherocytosis. SS and AA erythrocytes were also examined for changes in intracellular hemoglobin concentration of three different density fractions and with increasing duration of spin. The minimum force and duration of centrifugation required to impair water permeability were found to vary with the red cell type, the anticoagulant used (heparin or EDTA), the initial hematocrit of the sample centrifuged, as well as among the individual erythrocyte fractions within the same sample. When subjecting pathologic erythrocytes to high-speed centrifugation, the 125I-albumin dilution technique can be used to determine whether the centrifugation procedure has led to an artifactual red cell water loss and to correct for this when it does occur. An abnormal membrane susceptibility to mechanical stress was demonstrated in erythrocytes from patients with hereditary spherocytosis and several hemoglobinopathies.

Anemia, Sickle Cell↗
PubMed1991

Centrifugal ventricular assist devices.

A centrifugal ventricular assist device (VAD) uses centrifugal force to propel blood through nonocclusive pumpheads. Blood is diverted from either the right atrium or left atrium to a disposable pumphead. Then kinetic energy is added to blood contained within the VAD by rotating it at high speed. Centrifugal force generates energy which causes the blood to rise from the pumphead's base, thus forcing a return flow direction. Centrifugal VAD design has been used to sustain myocardial function when patients could not be weaned from cardiopulmonary bypass and as a bridge to transplantation. Interaction between centrifugal VAD and human hosts are complicated and multisystemic. Therefore a multidisciplinary VAD support team is most efficacious. A case study, nursing diagnoses, and care plan provide a comprehensive overview of nursing management for this patient population.

Centrifugation↗
PubMed1988

Malaria diagnosis by direct observation of centrifuged samples of blood.

We compared the effectiveness of malaria diagnosis by means of direct observation of centrifuged blood with that by conventional examination of Giemsa stained blood-films in a malaria clinic in Ethiopia. A commercially available, modified hematological apparatus (the QBC tube) was used for centrifugation. Red blood cells infected with diverse stages of Plasmodium falciparum and P. vivax are lighter than noninfected cells and somewhat heavier than granulocytes; thus they can readily be detected by direct inspection of UV-illuminated tubes. About 10% of infections diagnosed by direct centrifugal microscopy in a clinical setting were not detected by conventional examination of stained thick films. Diagnosis by direct centrifugation appears to be at least 8 times as sensitive as conventional microscopy when applied to serially diluted samples of malaria-infected blood. Superior sensitivity, together with the one step, solid state nature of the direct centrifugal procedure, provides important advantages for malaria diagnosis.

Acridine Orange↗
PubMedDate not supplied

Laboratory investigations on neuroparalytic accidents associated with suckling mouse brain rabies vaccine. III. -- Preservation of vaccine potency after elimination of murine brain myelin by centrifugation.

Myelin, which has been found in nine day-old mouse brain, was eliminated from weanling mouse brain suspensions by centrifugation at 17,000 g for 10 min, as demonstrated by electron microscopy and guinea-pig inoculation tests for encephalitogenic activity. This centrifugation procedure did not affect the potency of seven batches of suckling mouse brain vaccine, when centrifuged and non-centrifuged samples of the same batches were compared by a modified NIH potency test (t = 0.17). The present results with weanling mouse brain preparations suggest that centrifugation at 17,000 g be used in the preparation of rabies suckling mouse brain vaccine instead of the 1,900 g currently employed, which does not eliminate myelin. This new procedure would be expected to reduce the number of postvaccinal reactions which are attributable to the small amount of myelin which remains in vaccines prepared with new-born animal brains following the current procedure.

Animals↗
PubMed2000

Laboratory scaledown of protein purification processes involving fractional precipitation and centrifugal recovery.

The ability to predict the performance of large-scale processes is central to the rapid development of successful operations at the pilot and industrial scale. In this article, we examine the operation, at laboratory scale, of precipitation reactors and centrifuges for protein precipitate recovery and dewatering and how they might best mimic large-scale reactors and centrifuges, in this case, a pilot-scale batch stirred-tank reactor and a multichamber-bowl centrifuge. Novel approaches to bench-top centrifuge operation are provided, in particular with a view to delivery of material for subsequent high-resolution purification, which would be obtained at full pilot scale. Results are presented in terms of properties of the protein precipitates, the fraction of solids recovered, and the extent of dewatering achieved. Good agreement was obtained at bench scale (a 1000-fold scale down factor) for all of these parameters for pilot-scale, batch-feed operation. In addition, the methodology developed allows identification of the extent of break-up that occurs in continuous-feed centrifuges when processing shear-sensitive materials such as the protein precipitates studied here.

Alcohol Dehydrogenase↗
PubMed2001

Centrifugal pump and reduction of neurological risk in adult cardiac surgery.

This study was performed to assess if the kind of pump used for CPB (roller vs. centrifugal) can influence neurological outcomes of adult cardiac surgery patients. Between 1994 and 1998, 3438 patients underwent coronary and/or valve surgery at our hospital; of these, 1805 (52.5%) underwent surgery with the use of a centrifugal pump, and 1633 (47.5%) were operated with a roller pump. The effect of the type of the pump and of common preoperative and intraoperative risk factors for five different neurological outcomes (permanent neurological deficit, coma, delirium, transient neurological deficit, overall neurological complications) were assessed with univariate and multivariate analyses in the whole patients population, in patients > or = 75 years old and in patients with histories of previous neurological events. Centrifugal pump use was the only protective factor for perioperative permanent neurological deficit in multivariable models developed for the whole patient population and for patients > or = 75 years old. In addition, it resulted as the only protective factor for perioperative coma occurrence in multivariable models developed for patients > or = 75 years old, and for patients with histories of previous neurological events. The use of the centrifugal pump provided a risk reduction for the considered events ranging from 23 to 84%. Centrifugal pump use can be helpful in reducing the occurrence of some of the most feared neurological complications of adult cardiac surgery patients.

Aged↗