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PubMedDate not supplied

Percoll density gradient centrifugation and consecutive flow cytometry do not identify leukocytes and leukocyte subtypes in ejaculate specimens.

This paper describes an attempt to establish a new combined method of leukocyte analysis in human ejaculate by Percoll density gradient centrifugation and consecutive flow cytometry. As a first step, leukocyte separation was performed by Percoll density gradient centrifugation with consecutive enrichment of leukocytes, especially granulocytes, in the 40%/60% and 60%/80% Percoll interfaces. Then these fractions were stained with specific monoclonal antibodies and analysed in a Facscan flow cytometer. Flow cytometric analysis did not demonstrate identifiable leukocyte populations, indicating a questionable cross-reaction with spermatozoal elements. Therefore, the combined technique of Percoll density gradient centrifugation and flow cytometric analysis were considered unsuitable for clinical leukocyte determination.

Antibodies, Monoclonal↗

PubMedDate not supplied

Profiles of sperm morphology and motility after discontinuous multiple-step Percoll density gradient centrifugation.

Morphology and motility are important parameters for assessing the fertilizing capacity of spermatozoa. This investigation reports a systematic study on the profiles of these parameters after percoll gradient centrifugation. Spermatozoa from normal human semen were fractionated by discontinuous Percoll gradients (30%, 45%, 75%, 90%). Spermatozoa washed with Earle's balanced salt solution were used as a control. After centrifugation, sperm morphology was evaluated according to strict criteria; motility was assessed by a computer-assisted semen analysis system. The results showed that the percentage of spermatozoa with normal morphology increased, while those with severe morphological defects decreased, as the density of Percoll increased. The percentages of motile spermatozoa and hyperactivated spermatozoa, and the velocity and amplitude of lateral head displacement of the spermatozoa were significantly higher in the 75% and 90% Percoll fractions than that in the 30% and 45% ones, and in the control. These results demonstrated that Percoll density gradient centrifugation enriched spermatozoa in terms of morphology and motility.

Cell Separation↗

PubMed1980

The distribution of Mg2+ ATPase and its loss of specific activity upon gradient centrifugation of isolated chromaffin granules.

Preparations of purified chromaffin granules were subjected to isopycnic sucrose gradient centrifugation. Determination of Mg2+ ATPase activity and catecholamines showed that the distribution of ATPase almost parallelled the distribution of catecholamines. The distribution of ATPase was slightly shifted to lower densities and was suggested to be caused by the heterogeneity of the chromaffin granules. The results therefore provide evidence that ATPase is associated with chromaffin granules. Determination of the recovery of ATPase activity upon gradient centrifugation revealed losses of enzyme activity which were found to be proportional to the dilutions of the granule preparation subjected to gradient centrifugation.

Adenosine Triphosphatases↗

PubMed1995

Clinical experience with BioMedicus centrifugal ventricular support in 172 patients.

One hundred and seventy-two patients at The Methodist Hospital in Houston, Texas, were placed on BioMedicus centrifugal ventricular support. One hundred thirty-nine patients were male and 33 were female with a mean age of 59.7 years. Reasons for support were postcardiotomy cardiac failure (129 patients), cardiac allograft failure (17 patients), bridge to transplantation (10 patients), resuscitation (7 patients), postpercutaneous transluminal coronary angioplasty emergent (2 patients), and other (7 patients). Support was by left ventricular assist device in 108 patients, right ventricular assist device in 20 patients, and biventricular assist device in 44 patients. Eighty-four patients (48.8%) were weaned from the ventricular assist device, and 88 patients (51.2%) were not weaned. Thirty-four patients (20.0%) were discharged from the hospital. Complications included coagulopathy, renal insufficiency/failure, respiratory insufficiency/failure, neurological deficits, sepsis, arrhythmias, and device-related complications. Overall causes of death were ventricular failure (55.1%), triage (13.0%), arrhythmias (9.4%), graft failure (5.9%), coagulopathy (4.3%), sepsis syndrome (2.9%), device-related (0.7%), and other (0.7%). BioMedicus centrifugal ventricular support can be implemented rapidly and easily. Device-related complications are few (1.2%), and it is relatively inexpensive when compared with other ventricular assist systems. This series demonstrates that a substantial number of patients may benefit from temporary centrifugal ventricular support.

Adult↗

PubMed1995

Improving clinical outcome with centrifugal mechanical assist for postcardiotomy ventricular failure.

Between October 1986 and May 1994, 65 patients undergoing cardiac surgery required centrifugal mechanical assist devices to separate from cardiopulmonary bypass. This experience was arbitrarily divided into early (n = 33) and recent (n = 32) groups for the purpose of comparing trends in morbidity and mortality. The incidence of mechanical assist application decreased from 2.19% in the early group to 0.96% in the recent group (p < 0.0001). Ability to wean patients from centrifugal assist increased from 33% in the early group to 53% in the more recent group, and hospital survival increased from 15 to 28%. The median chest tube drainage during the first 24 h decreased from 3,245 ml to 1,535 ml, and the incidence of renal failure decreased from 39.4% and 18.8% in the more recent group. Clinically relevant improvement in patient outcome following application of centrifugal mechanical assist for postcardiotomy ventricular failure is being observed.

Adult↗

PubMed1996

A new magnetically suspended centrifugal pump: in vitro and preliminary in vivo assessment.

To overcome problems derived from the shaft within the conventional centrifugal pump, we have been developing a new centrifugal pump, namely a magnetically suspended centrifugal pump (MSCP), which has no shaft and operates as a noncontacting and bearingless pump. The impeller is suspended magnetically between the magnetic bearing and the driving motor. Hemolysis tests were performed in comparison with the Biopump (BP80, BioMedicus). The index of hemolysis (IH) was significantly lower in the MSCP than in the Biopump. In addition, a smaller gap in the MSCP induced lower hemolysis. In preliminary studies using mongrel dogs, the layer of thrombus adherent to the impeller was observed in a few hours, which impaired the pumping efficiency. However, by using an impeller coated with silicone, no aggregations of platelets or fibrin on the impeller were observed in 24 h of continuous pumping. In conclusion, the MSCP had a gentler influence on blood cells than the Biopump, and the impeller coated with silicone may contribute to the long-term pumping of the MSCP.

Animals↗

PubMed1997

In vitro investigation of the St. Jude Medical Isoflow centrifugal pump: flow visualization and hemolysis studies.

Centrifugal blood pumps have become valuable therapeutic tools for cardiopulmonary bypass surgery. In addition, surgeons have used them as temporary ventricular assist devices, and this type of pump is also being developed for use as a permanent assist device and total artificial heart. However, centrifugal pumps create flow patterns that are significantly different from those the blood experiences physiologically. The St. Jude Medical Isoflow centrifugal pump has been used clinically during cardiopulmonary bypass surgery, yet no experimental results have been reported that describe the flow patterns within this pump or that quantify the hemolysis generated over a range of operating conditions. The purpose of this study was to investigate the flow patterns and hemolysis during 4 operating conditions. The experimental operating conditions included the design condition (6 L/min, 2,500 rpm, 350 mm Hg), a high flow condition (10 L/min, 2,500 rpm, 330 mm Hg), a low flow condition (2 L/min, 2,500 rpm, 370 mm Hg), and a near surge condition (2 L/min, 3,000 rpm, 550 mm Hg). The flow visualization results demonstrated that the flow within the impeller was well aligned with the impeller blades except near the inlet at the high flow condition. In contrast, the flow through the outlet was well aligned at the high flow condition while there was evidence of particle impact at the design condition, and the flow was disturbed at the low flow and near surge conditions. The indices of hemolysis (IH) for the 3 operating conditions at 2,500 rpm were 0.0082 +/- 0.0026 (mean +/- SD) for the design condition, 0.0035 +/- 0.0014 for the high flow condition, and 0.0326 +/- 0.0050 for the low flow condition. The indices for high and low flow were significantly different from that for the design condition (p < 0.05). The IH for the near surge condition (0.0748 +/- 0.0039) was significantly higher than that for all other conditions (p < 0.05). In addition to describing the flow patterns within the Isoflow, this study independently validated St. Jude Medical's reported IH at the design condition and showed how that IH significantly changed based on operating conditions.

Blood Flow Velocity↗

PubMed1997

Vibration assessment for thrombus formation in the centrifugal pump.

To clarify the correlation of vibration and thrombus formation inside a rotary blood pump, 40 preliminary vibration studies were performed on pivot bearing centrifugal pumps. No such studies were found in the literature. The primary data acquisition equipment included an accelerometer (Isotron PE accelerometer, ENDEVCO, San Juan Capistrano, CA, U.S.A.), digitizing oscilloscope (TDS 420, Tektronix Inc., Pittsfield, MA, U.S.A.), and pivot bearing centrifugal pumps. The pump impeller was coupled magnetically to the driver magnet. The accelerometer was mounted on the top of the pump casing to sense radial and axial accelerations. To simulate the 3 common areas of thrombus formation, a piece of silicone rubber was attached to each of the following 3 locations as described: a circular shape on the center bottom of the impeller (CI), an eccentric shape on the bottom of the impeller (EI), and a circular shape on the center bottom casing (CC). A fast Fourier transform (FFT) method at 5 L/min against 100 mm Hg, with a pump rotating speed of 1,600 rpm was used. The frequency response of the vibration sensors used spans of 40 Hz to 2 kHz. The frequency domain was already integrated into the oscilloscope, allowing for comparison of the vibration results. The area of frequency domain at a radial direction was 206 +/- 12.7 mVHz in CI, 239.5 +/- 12.1 mVHz in EI, 365 +/- 12.9 mVHz in CC, and 163 +/- 7.9 mVHz in the control (control vs. CI p = 0.07, control vs. EI p < 0.001, control vs. CC p < 0.001, EI vs. CC p < 0.001, CI vs. CC p < 0.001). Three types of imitation thrombus formations were roughly distinguishable. These results suggested the possibility of detecting thrombus formation using vibration signals, and these studies revealed the usefulness of vibration monitoring to detect thrombus formation in a centrifugal pump.

Analysis of Variance↗

PubMed1997

Platelet damage caused by the centrifugal pump: in vitro evaluation by measuring the release of alpha-granule packing proteins.

Platelets are more vulnerable to damage than erythrocytes because platelets are easily activated by contact with extracorporeal circuits and by exposure to shear forces. However, the degree of platelet damage caused by centrifugal pumps is unclear. To evaluate platelet damage in different pumping conditions, the rates of increase for specific proteins in platelet alpha-granules, beta-thromboglobulin (beta-TG), and platelet factor 4 (PF-4) were measured in both in vitro simulated left ventricular assist device (LVAD) and cardiopulmonary bypass (CPB) conditions and compared with the erythrocyte trauma. A flow of 5.0 L/min with deltaP of 100 mm Hg for LVAD (low pressure head condition) and a flow of 5.0 L/min with deltaP of 350 mm Hg for CPB (high pressure head condition) were investigated. Each condition was tested 4 times for 3 h in a mock circuit with a Capiox (Terumo, Tokyo, Japan) centrifugal pump using fresh human blood. Blood was sampled at 1 h intervals, measuring plasma free hemoglobin (fHb), beta-TG, and PF-4. To evaluate the degree of damage, the rates of increase of fHb, beta-TG, and PF-4 were calculated for each condition as deltafHb/deltaN, deltabeta-TG/deltaN, and deltaPF-4/deltaN where deltafHb is the increase in plasma free hemoglobin, deltabeta-TG is the increase in beta-TG, deltaPF-4 is the increase in PF-4, and deltaN is the increase in the passing number. The passing number is defined in the following equation: N = Qt/V where t is the time, V is the priming volume, and Q is the flow rate. There was no significant difference between the 2 conditions (low pressure head condition versus high pressure head condition) in the rate of increase of fHb (0.0035+/-0.0004 vs. 0.0034+/-0.0010 g/100 L, NS). Contrary to this, the rates of increase for specific proteins in platelet alpha-granules in the high pressure head condition demonstrated a significantly higher rate of increase than in the low pressure head condition. The mean rate of increase for beta-TG in the low pressure head condition was 0.22+/-0.03 ng/ml and in the high pressure head condition was 0.51+/-0.05 ng/ml (p < 0.05). The rate of increase for PF-4 in the low pressure head condition was 0.11+/-0.02 ng/ml and in the high pressure head condition was 0.30+/-0.06 ng/ml (p < 0.05). These results suggest that measurements of beta-TG and PF-4 may be more sensitive parameters than hemolysis for evaluating blood cell trauma and that platelets are more vulnerable to mechanical damage by a centrifugal pump than erythrocytes.

Blood Platelets↗

PubMed1997

The method for keeping low perfusion flow weaning from centrifugal pumps: an evaluation of hemolysis in the circulator units.

Currently, the clinical application of extracorporeal and assisted circulation using centrifugal pumps is becoming popular. However, a problem in the use of centrifugal pumps is the difficulty in controlling the flow rate when it is a low range. Although we have been controlling the flow rate with a control unit that simply squeezes the blood outlet tube from the outside, hemolysis resulting from turbulent flow in the tube was discovered. Therefore, this control procedure was evaluated experimentally. Two groups were tested as follows: in Group A (n = 10), the tube was squeezed from the outside using the control unit, and in Group B (n = 10), no control unit was used. Blood in each circuit was circulated at 1,500 rpm, and the flow rate was controlled to 250 ml/min using the control unit for Group A. Time-course changes of lactate dehydrogenase (LDH) and free hemoglobin in the blood were measured during 8 h of circulation. There were no significant differences between the 2 groups in the mean values of LDH and free hemoglobin. Therefore, the present low flow rate control provided by the control unit makes it appear to be a simple and useful apparatus for controlling blood circulation using centrifugal pumps.

Analysis of Variance↗

PubMed1997

A new blood pump for cardiopulmonary bypass: the HiFlow centrifugal pump.

Centrifugal blood pumps are considered to be generally superior to the traditionally used roller pumps in cardiopulmonary bypass. In our institute a new lightweight centrifugal sealless blood pump with a unique spherical thrust bearing and with a magnetic coupling was developed, the HiFlow. The small design makes the pump suitable for applications in complex devices or close to a patient. Hemolysis tests were carried out in which the BioMedicus pump BP-80 and a roller pump were used as reference. The centrifugal pump HiFlow showed the least blood trauma within the group of investigated pumps. In summary, the HiFlow pump concept with its low priming volume and limited contact surfaces shows great potential for clinical applications in cardiopulmonary bypass. Also, the possibility of using the pump as a short-term assist device with an option of a pulsatile driving mode was demonstrated.

Animals↗

PubMed2005

Design considerations of volute geometry of a centrifugal blood pump.

This article compares two different design techniques that are conventionally used in the design of volutes for centrifugal pumps. The imbalanced forces due to the geometry of the volute need to be taken into consideration especially in centrifugal blood pumps with magnetically suspended impeller. A reduction of these forces can reduce the instability of the impeller motion as well as the power needed to counteract its influence. Volutes using the constant angular momentum (CAM) and the constant mean velocity (CMV) methods were developed and modeled numerically. The computational results on the effect of volute geometry on the performance of a centrifugal blood pump impeller for six different volutes are presented here. For volutes designed using the CAM method, model B (volute expansion angle of 3 degrees ) had the lowest radial force of 0.26 N while the pressure head generated was 12,900 Pa. For volutes designed using the CMV method, model F (1.6 m/s) had the lowest imbalanced force of 0.45 N. However, the pressure developed by this pump was also one of the lowest at 10,652 Pa. Furthermore, when the peak scalar stresses and the mean exposure time of particles for all designs were determined using Lagrangian particle tracking method, it was observed that in general, the peak scalar stresses in CAM designed volutes are lower than those designed using CMV method. The mean exposure time of particles in the pump ranged from 400 to 500 ms. The simulation results showed that the volute designed using CAM method was superior to that of a CMV volute in terms of the magnitude of the radial force and the peak scalar stresses for the same pressure head generated. Results show that the design of volutes for blood pumps should go beyond conventional empirical methods to obtain optimal results.

Centrifugation↗

PubMed2006

Computational fluid dynamics analysis of the pediatric tiny centrifugal blood pump (TinyPump).

We have developed a tiny rotary centrifugal blood pump for the purpose of supporting circulation of children and infants. The pump is designed to provide a flow of 0.1-4.0 L/min against a head pressure of 50-120 mm Hg. The diameter of the impeller is 30 mm with six straight vanes. The impeller is supported by a hydrodynamic bearing at its center and rotated with a radial coupled magnetic driver. The bearing that supports rotation of the impeller of the tiny centrifugal blood pump is very critical to achieve durability, and clot-free and antihemolytic performance. In this study, computational fluid dynamics (CFD) analysis was performed to quantify the secondary flow through the hydrodynamic bearing at the center of the impeller and investigated the effects of bearing clearance on shear stress to optimize hemolytic performance of the pump. Two types of bearing clearance (0.1 and 0.2 mm) were studied. The wall shear stress of the 0.1-mm bearing clearance was lower than that of 0.2-mm bearing clearance at 2 L/min and 3000 rpm. This was because the axial component of the shear rate significantly decreased due to the narrower clearance even though the circumferential component of the shear rate increased. Hemolysis tests showed that the normalized index of hemolysis was reduced to 0.0076 g/100 L when the bearing clearance was reduced to 0.1 mm. It was found that the CFD prediction supported the experimental trend. The CFD is a useful tool for optimization of the hydrodynamic bearing design of the centrifugal rotary blood pump to optimize the performance of the pump in terms of mechanical effect on blood cell elements, durability of the bearing, and antithrombogenic performance.

Biomedical Engineering↗

PubMed2004

Compatibility testing without a centrifuge: the slide Polybrene method.

BACKGROUND: A simple and rapid slide Polybrene method (SP) for pretransfusion compatibility testing is described. SP is particularly suitable for use in developing countries where, due to limited resources, centrifuges and biologic reagents may not be readily available. STUDY DESIGN AND METHODS: The original manual Polybrene method (MP) was modified for use on glass microscope slides, eliminating the need for test tubes and centrifugation. The sensitivity of SP for detecting alloantibodies to RBC antigens was compared with that of MP and the IAT. RESULTS: Both SP and MP were more sensitive than the IAT for detecting anti-E. SP detected 21 of 23 examples of anti'-Mi(a)' and 7 of 8 examples of anti-E. Kidd and Diego system antibodies were also readily detectable by SP, although the reactions were weaker than those observed with both MP and IAT. However, both SP and MP failed to detect some examples of antibodies to Kell system antigens. CONCLUSIONS: SP is an acceptable method for compatibility testing in developing countries, particularly in populations where the frequency of K is low (e.g., southeast Asia). The reagents are inexpensive and can be prepared in-house. SP is simple to use, does not require a centrifuge, and can be performed by personnel with minimal training.

Antibodies↗

PubMed1986

Glycolytic activity in human red cell populations separated by a combination of density and counterflow centrifugation. Evidence for an improved separation of red cells according to age.

Human red blood cells were separated by a discontinuous Percoll density gradient into fractions of increasing density. Red cells comprising the lowest and highest density fractions, respectively, were subsequently separated according to mean cell volume (MCV) by means of counterflow centrifugation. The activities of 4 red cell age-dependent enzymes (hexokinase (HK), pyruvate kinase (PK), glucose-6-phosphate dehydrogenase (G6PD) and aspartate aminotransferase (ASAT) were highest in the red cell fraction with low density/large MCV, although the difference from red cell enzyme activities in the total low density fraction was not significant. These 4 enzyme activities were lowest in the fraction of red cells with high density/small MCV. The relative activities of the enzymes in the high density/small MCV fraction, as compared to the unseparated cell population, were: HK (58%), PK (49%), G6PD (53%) and ASAT (28%). These activities were all significantly lower than those measured in the total high density red cell fraction. The rates of lactate production in the low density/large MCV cells (0.89 +/- 0.15 mumol X min-1 X 10(-11) cells) is approximately 3-fold higher than in high density/small MCV cells (0.33 +/- 0.03 mumol X min-1 X 10(-11) cells). This latter value is 1.8-fold lower than the rate of lactate production in the total high density red cell fraction (0.59 +/- 0.14 mumol X min-1 X 10(-11) cells) and is, in contrast to lactate production in other density/size fractions, insensitive to phosphate as a metabolic stimulus. It is argued that the combination of density gradient and counter-flow centrifugation offers a greater potential for obtaining an old red cell population than classical red cell density centrifugation alone.

Cell Separation↗

PubMed1997

Spin doctors: new innovations for centrifugal apheresis.

The preparation of plasma from blood has a long history dating back to the early 1900s when the concept of blood washing replaced the traditional blood letting. Over the next 57 years landmark discoveries such as centrifugal and membrane filtration systems led to different and rapid plasma, solute, and cell separation. These were not singular events but rather events influenced by the converging chemical, physiological, and engineering advances that have characterized the latter half of the 20th century. These events have led to entire new fields of biomedical research. The biotechnology for on-line plasma separation and plasma treatment has opened a new era, expanding the application of extracorporeal technology to modern therapeutic medicine. The association of biochemical or cellular abnormalities with various disease states provides the rationale for therapeutic plasma exchange (the removal of large amounts of patient's plasma, alone or with replacement with crystalloid) and therapeutic cytopheresis (removal of cellular elements). The purpose of this review is to provide a historical picture of the innovative ideas of the spin doctors and their devices, which predate the centrifugal blood and cell separators commonplace to any hospital or blood bank worldwide. The emphasis is to define the historical events and their impacts on the development of centrifugal devices and apheresis technologies.

Blood Component Removal↗

PubMed1998

Centrifugal leukocyte apheresis for ulcerative colitis.

Corticosteroids are effective in bringing about clinical remission for ulcerative colitis. However, relapsed cases are frequently refractory to corticosteroids. In addition, the long-term use of corticosteroids often causes serious side effects. We have already reported that leukocyte apheresis using a centrifugal procedure was effective for patients with corticosteroid-resistant ulcerative colitis after conducting a pilot study in 14 patients. In the present paper, the clinical efficacy of leukocyte apheresis using a centrifugal procedure was evaluated again for corticosteroid-resistant ulcerative colitis. Twenty-three patients with corticosteroid-resistant severely active ulcerative colitis were treated by centrifugal leukocyte apheresis. Eighteen patients (78.3%) achieved clinical remission which was evaluated by the clinical activity index within 4 weeks after apheresis. Both colonoscopic and histological examinations confirmed the beneficial effect of this procedure. No significant side effects were noticed throughout the therapy.

Adult↗

PubMed1999

Human ocular counterrolling during roll-tilt and centrifugation.

To test a hypothesis about how otoliths resolve roll-tilts from translations, we measured human ocular torsion position [ocular counterrolling (OCR)] to maintained linear acceleration stimuli. All subjects (n = 8) were tested in two conditions where the same magnitude of shear along an interaural axis was generated in one of two ways: either by roll-tilt on a tilt-chair in a 1-g environment, or by centripetal linear acceleration during constant velocity rotation 1 m from the axis of rotation on a fixed-chair human centrifuge. The interaural shear to the otoliths was the same for these two conditions, but the dorsoventral shear was different and for all eight subjects the OCR on the centrifuge was significantly greater than the torsion on the tilt-chair, although the resultant angle was in fact smaller on the centrifuge than on the tilt-chair. The results confirm that dorsoventral shear is important for determining OCR. The otoliths may resolve potential stimulus ambiguities between tilts and translations by virtue of the different patterns of interaural and dorsoventral shear that these stimuli generate.

Acceleration↗