Red Cell Separation Research Articles

Cell Salvage Using the Autotransfusion Device CATSmart®: A Randomized Controlled Bicentric Trial Evaluating the Quality of Two New Flex Wash Programs

Background: The use of cell salvage and autologous blood transfusion is an important and widespread method of blood conservation during surgeries with expected high blood loss. The continuous autotransfusion device CATSmart^® (Fresenius Kabi, Germany) contains two new washing programs on the device called Flex wash 3 and Flex wash 5. To the best of our knowledge, there are no published clinical data regarding the performance of the two new washing programs. Methods: In total, 69 patients undergoing cardiac or orthopedic surgery were included in this randomized, controlled, bicentric trial to validate the red cell separation process and washout quality of Flex wash 3 compared to Flex wash 5. After washing, the primary quality target was to determine hematocrit value, recovery rate, albumin, and total protein elimination rate in the packed red cells (PRCs). The secondary objective was to assess the elimination of heparin by measuring the factor anti-Xa activity by a 1- and 2-stage assay in PRC after washing. Results: In the whole cohort of patients, hematocrit was 16.00% [9.15%; 21.30%] (median [Q1; Q3]) in the wound blood and 69.90% [51.10%; 80.90%] in the PRC resulting in a recovery rate of 63.92% [47.06%; 88.13%]. The albumin elimination rate was 98.77% [97.94%; 99.27%], and the total protein elimination rate was 98.85% [97.76%; 99.42%]. The heparin elimination rate was 99.95% [99.90%; 99.97%] in the 1-stage assay and 99.70% [99.41%; 99.87%] in the 2-stage assay. There was no difference between Flex wash 3 and Flex wash 5 washing procedure regarding the recovery rate 63.75% [46.64%; 78.65%] versus 67.89% [47.20%; 92.69%] (p = 0.85), albumin elimination rate 98.74% [97.67%; 99.27%] versus 98.78% [98.10%; 99.28%] (p = 0.97), protein elimination rate 98.79% [97.94%; 99.47%] versus 98.92% [97.58%; 99.42%] (p = 0.88), and anti-Xa elimination rate in the 1-stage assay 99.94% [99.79%; 99.97%] versus 99.95% [99.92%; 99.97%] (p = 0.24) and in 2-stage assay 99.66% [99.20%; 99.86%] versus 99.77% [99.47%; 99.90%] (p = 0.23). Conclusions: The two new washing procedures, Flex wash 3 and Flex wash 5, enable sufficient and comparable red cell separation and washout quality of albumin, total protein, as well as heparin.

Cell salvage using the continuous autotransfusion device CATSmart \u2013 an observational bicenter technical evaluation

BackgroundThe use of cell salvage and autologous blood transfusion has become an important method of blood conservation. So far, there are no clinical data about the performance of the continuous autotransfusion device CATSmart.MethodsIn total, 74 patients undergoing either cardiac or orthopedic surgery were included in this prospective, bicenter and observational technical evaluation to validate red cell separation process and washout quality of CATSmart. The target of red cell separation process was defined as a hematocrit value in the packed red cell unit of 55–75% and of washout quality of 80–100% removal ratio.ResultsHematocrit values measured by CATSmart and laboratory analysis were 78.5% [71.3%; 84.0%] and 73.7% [67.5%; 75.5%], respectively. Removal ratios for platelets 94.7% [88.2%; 96.7%], free hemoglobin 89.3% [85.2%; 94.9%], albumin 97.9% [96.6%; 98.5%], heparin 99.9% [99.9%; 100.0%], and potassium 92.5% [90.8%; 95.0%] were within the target range while removal of white blood cells was slightly worse 72.4% [57.9%; 87.3%].ConclusionThe new autotransfusion device enables sufficient red cell separation and washout quality.

Paper Diagnostic for Instantaneous Blood Typing

Agglutinated blood transports differently onto paper than stable blood with well dispersed red cells. This difference was investigated to develop instantaneous blood typing tests using specific antibody-antigen interactions to trigger blood agglutination. Two series of experiments were performed. The first related the level of agglutination and the fluidic properties of blood on its transport in paper. Blood samples were mixed at different ratios with specific and nonspecific antibodies; a droplet of each mixture was deposited onto a filter paper strip, and the kinetics of wicking and red cell separation were measured. Agglutinated blood phase separated, with the red blood cells (RBC) forming a distinct spot upon contact with paper while the plasma wicked; in contrast, stable blood suspensions wicked uniformly. The second study analyzed the wicking and the chromatographic separation of droplets of blood deposited onto paper strips pretreated with specific and nonspecific antibodies. Drastic differences in transport occurred. Blood agglutinated by interaction with one of its specific antibodies phase separated, causing a chromatographic separation. The red cells wicked very little while the plasma wicked at a faster rate than the original blood sample. Blood agglutination and wicking in paper followed the concepts of colloids chemistry. The immunoglobin M antibodies agglutinated the red blood cells by polymer bridging, upon selective adsorption on the specific antigen at their surface. The transport kinetics was viscosity controlled, with the viscosity of red cells drastically increasing upon blood agglutination. Three arm prototypes were investigated for single-step blood typing.

Adenosine A3 receptor activation modulates the capillary endothelial glycocalyx.

The endothelial glycocalyx is a dynamic extracellular matrix composed of cell surface proteoglycans, glycoproteins, and adsorbed serum proteins that has been implicated in the regulation and modulation of capillary tube hematocrit, permeability, and hemostasis. High tissue adenosine levels have been shown to adversely affect microvascular function and tissue survival after an ischemic episode, and previous work in this laboratory has shown that adenosine causes arteriolar constriction and degranulation of mast cells via the A3 receptor (A3AR). We hypothesized that adenosine exerts at least part of its effect through modification of the glycocalyx via the A3AR. We used an in vivo cremaster model (hamster and mouse) in which circulating plasma was labeled with a 70-kDa FITC-dextran, and the capillaries were examined before and after superfusion with varying concentrations of adenosine (or other vasoactive molecules). Measurements of the dextran exclusion from an endothelial cell surface layer and red cell separation from the endothelial cell surface were made for up to 30 minutes. Our data indicate that adenosine causes a rapid and profound decrease in the ability of the glycocalyx to exclude dextran but only affects red blood cell exclusion at pharmacological levels. Knockout mice deficient in the A3AR were completely protected from glycocalyx changes attributable to adenosine. These data show a potential link between a known vasoactive tissue metabolite, adenosine, and regulation of the glycocalyx, which may be important during (patho)physiological changes in microvascular function during inflammatory insults.

Comparison of the effects of red cell separation and ultrafiltration on heparin concentration during pediatric cardiac surgery

Objective: To determine the effects of red cell separation and ultrafiltration on heparin concentration. Design: Prospective study. Setting: University-affiliated, pediatric medical center. Participants: Thirty-one children undergoing cardiac surgery. Interventions: Blood sampled for heparin concentration and coagulation tests. Measurements and Main Results: Thirteen infants underwent modified veno-venous ultrafiltration (UF) after cardiopulmonary bypass (CPB). In addition, residual blood in the CPB circuit was hemoconcentrated by UF and reinfused (UF group). Heparin concentration increased from 2.0 ± 0.6 to 2.5 ± 0.8 U/mL, following modified UF; while activated coagulation time (ACT) decreased from 701 ± 177 to 627 ± 107 seconds. Heparin concentration of CPB circuit residual increased from 1.9 ± 0.7 to 3.1 ± 1.0 U/mL. In 18 children (older than 1 year old), the residual blood in the CPB circuit was hemoconcentrated by cell separation (CS) and reinfused (CS group). Heparin concentration of CPB circuit residual decreased from 2.6 ± 0.6 to 0.3 ± 0.2 U/mL. After reinfusion, patient heparin concentration remained unchanged at <0.05 U/mL. Thrombin time increased from 28 ± 6 to 48 ± 29 seconds and did not correlate with H. Conclusions: The plasma concentration of heparin increased after veno-venous modified UF of the patient. Heparin concentration also increased after UF of residual CPB circuit blood. In contrast, circuit blood hemoconcentrated by CS contained minimal heparin, and, when infused, did not increase patient's heparin concentration. ACT and thrombin time did not correlate with heparin concentration.

An examination of the contribution of red cell spacing to the uniformity of oxygen flux at the capillary wall

It is generally assumed that capillary blood is homogeneous for O 2 supply and that red cells can provide a constant, uniform flux of O 2 out of the capillary regardless of the spacing between cells. Using a simplified model of red cells moving through a capillary in skeletal muscle, an approximate analysis is developed to study the effect of red cell spacing on the ability of erythrocytes to provide a constant, uniform flux of O 2 at the capillary wall. The results suggest the existence of a critical red cell separation distance above which the flux of O 2 at the capillary wall between red cells cannot remain uniform and the capillary blood is no longer homogeneous for O 2 supply. In resting muscle the predicted critical separation distance is greater than four cell lengths. During maximal O 2 consumption, the critical separation distance predicted by the model is one cell length. These predictions agree closely with in vivo observations of red cell spacing. The total red cell flux through a capillary is determined not only by red cell spacing (hematocrit) but also by erythrocyte velocity; a simple example is given which suggests that changes in each of these variables are not equivalent in maintaining a constant and uniform flux of O 2 at the capillary wall.

Ex vivo selective isolation of young red blood cells using the IBM-2991 cell washer

Transfusion-induced hemochromatosis is a major complication in the therapy of severe chronic anemia. Improvement of transfused cell survival with a reduction in transfusion frequency is one possible approach to this problem. Using continuous-flow centrifugation (CFC), young red blood cells (YRBC) with enhanced in vivo survival have been isolated, but the expense and donor time required with this technique prohibit its widespread use for patient support. We studied the use of the IBM 2991 cell washer (CW) to isolate YRBC ex vivo from previously collected donor blood. Age-dependent red cell separation could be achieved using this instrumentation. Autologous mean red cell half-life (RBC-T50) (n = 9) for the younger cell fractions was 43.9 +/- 7.8 days compared to 34.7 +/- 5.8 days for the older cell fractions (n = 6, p less than 0.05). Paired measurement of RBC-T50 for young and old fractions in three donors showed an average survival increase of 41% for the YRBC. Adequate quantities of YRBC with enhanced survival can be obtained with less cost and less donor stress using the CW system compared to CFC. This approach could improve the management of patients with chronic transfusion requirements and merits further examination.

Ex Vivo Selective Isolation of Young Red Blood Cells Using the IBM-2991 Cell Washer

Transfusion-induced hemochromatosis is a major complication in the therapy of severe chronic anemia. Improvement of transfused cell survival with a reduction in transfusion frequency is one possible approach to this problem. Using continuous-flow centrifugation (CFC), young red blood cells (YRBC) with enhanced in vivo survival have been isolated, but the expense and donor time required with this technique prohibit its widespread use for patient support. We studied the use of the IBM 2991 cell washer (CW) to isolate YRBC ex vivo from previously collected donor blood. Age-dependent red cell separation could be achieved using this instrumentation. Autologous mean red cell half-life (RBC-T50) (n = 9) for the younger cell fractions was 43.9 +/- 7.8 days compared to 34.7 +/- 5.8 days for the older cell fractions (n = 6, p less than 0.05). Paired measurement of RBC-T50 for young and old fractions in three donors showed an average survival increase of 41% for the YRBC. Adequate quantities of YRBC with enhanced survival can be obtained with less cost and less donor stress using the CW system compared to CFC. This approach could improve the management of patients with chronic transfusion requirements and merits further examination.

Human erythrocyte separation according to age on a discontinuous "Percoll" density gradient.

It is well known that during ageing, erythrocyte density increases [l-3]. Density gradient centrifugation is the technique most generally used for fractionating erythrocytes of different mean age. Materials employed to construct density gradients, such as bovine serum albumin (BSA) [4], Stractan II [5], Ficoll [6] and Dextran [7], have many disadvantages. The development of Percoll (colloidal silica particles coated with polyvinylpyrrolidone) has overcome these problems. Owing to its low viscosity, low osmotic pressure and non-toxicity this material can be easily adjusted to physiological conditions. Percoll methods allow separation of erythrocytes on discontinuous gradients [&lo], linear gradients [ 111, or self-generated gradients [ 121. In this work, red cell separation in a discontinuous gradient similar to that reported by Alderman et al. [9] is described. To obtain Percoll with the closest approximation to physiological conditions, a suitable modification of Rennie’s buffer system [ 1 l] has been used. Separated red cell populations have been tested for cell recoveries, reticulocyte count and some age-dependent erythrocyte enzyme activities (pyruvate kinase, hexokinase, glucose-6-phosphate dehydrogenase, aldolase) and indices (mean cell volume and red cell volume distribution width index, mean cell haemoglobin concentration). Two non age-dependent measurements were also made (phosphoglycerate kinase and mean cell haemoglobin). The effectiveness and reliability of the method have been evaluated.

Red cell separation in the mesenteric capillary network.

Red cell separation in the mesenteric capillary network.

Magnetoelectrophoresis.

The theory of magnetoelectrophoresis of erythrocytes is described and a system of red cell separation from plasma is presented in detail. The apparatus has been designed and built and tested to demonstrate its effectiveness. The steps necessary to make this process applicable to the routine donor room are described.