Cell Transit Analyzer Research Articles

Use of cell transit analyzer to measure the film thickness around red blood cells

Use of cell transit analyzer to measure the film thickness around red blood cells

Effects of calcium permeabilization on RBC rheologic behavior.

Although many RBC rheological properties have been previously described in detail, the biochemical mechanisms leading to premature destruction of red blood cells are less clear. However, several biochemical processes have been suggested as possible mechanisms for membrane structural alterations (e.g., crosslinking of membrane proteins, oxidant damage, binding of cytoplasmic proteins, and altered intracellular ion composition). We have carried out a series of studies aimed at evaluating the effects of calcium-regulated membrane-bound hemoglobin (Hbm) on RBC and derived ghost rheologic behavior. Intracellular calcium was elevated by 10 microM A23187, with cell deformability determined via the Cell Transit Analyzer (CTA). Our results indicate: 1) Linear, positive correlations between Hbm and average RBC rigidity and 2) a marked influence of heterogeneous calcium concentration on both Hbm and rheologic properties for various subpopulation. These findings therefore suggest the importance of hemoglobin-membrane interactions as a determinant of erythrocyte deformability, and may be relevant to RBC aging as well as to diseases such as sickle cell anemia, hereditary spherocytosis and thalassemia.

Assessment of red blood cell deformability in healthy adult subjects: Influence of sex, age, cigarette smoking, menstrual cycle and oral contraceptives

To assess the possible influence of age, gender, cigarette smoking, menstrual cycle and oral contraceptives on red blood cell (RBC) deformability, cell transit parameters for one-hundred-ninety-four healthy adult subjects (seventy-nine males and one-hundred-fiveteen females) have been measured by filtration with the Cell Transit Analyser (CTA). The CTA provides the distribution of cell transit times of 5000 red blood cells, the mean transit time of the RBC population and different percentiles such as p50, p75, p90 and p95. First, correlations between and mean cell volume (MCV) were calculated for the entire population. Secondly, the influence of sex, age, cigarette smoking, menstrual cycle and oral contraceptives on the RBC mean transit time was evaluated. For RBC with 80 fl and MCV was found (r = 0.41; P < 0.001). No effect of age, cigarette smoking and oral contraceptives was found. Nevertheless, influence of sex and menstrual cycle were demonstrated. A significant increase of the CTA parameters measured in the female population with respect to the male population and during menstruation, preovulation and post-ovulation periods was observed. During ovulation, the CTA parameters are comparable to the same parameters in males.

Laser-assisted optical rotational cell analyser (L.O.R.C.A.); II. Red blood cell deformability: Elongation index versus cell transit time

Red blood cell deformability was measured under normal and experimental conditions with the Laser-assisted Optical Rotational Cell Analyser (LORCA). Results were compared to simultaneously performed filtration measurements using the Cell Transit Analyser (CTA). The deformability parameters of the two methods were Elongation Index (EI) and mean Cell Transit Time (CIT), respectively. Aside from instrumental and intra-individual reproducibility, normal values, the effect of sample aging and of changing medium osmolality are reported. Furthermore, the respective sensitivities were determined using red blood cells with slightly impaired deformability. Both heat treatment and incubation in the presence of low concentrations of glutaraldehyde are used to manipulate the deformability of the entire red cell population. It is concluded that minor defects in red blood cell deformability are detected more easily by the laser technique than by filtration. Even when the absolute differences in EI are small, they can be significant due to the very high reproducibility of the ektacytometric technique. Differentiation between a decreased deformability of the entire RBC population and the presence of a small (i.e., 5% or higher) subpopulation of rigid cells can be achieved with the LORCA by combining the measured EI-values with visual inspection of the diffraction pattern. Finally, no significant correlation was found between the results of both techniques applied to blood from 525 patients with various diseases.

How to detect alterations of the red blood cell mechanical properties from cell transit analyser filtration experiments

How to detect alterations of the red blood cell mechanical properties from cell transit analyser filtration experiments

Influence of calcium permeabilization and membrane-attached hemoglobin on erythrocyte deformability.

The present study was designed to evaluate the influence of intracellular calcium [Ca]i regulated membrane attached hemoglobin (Hbm) on the deformability of human RBC and ghosts. [Ca]i of RBC was elevated via the ionophore A23187 (10 microM); the deformability of RBC and resealed ghosts was determined via measuring RBC and ghost transit times through 5 microns diameter pores with the Cell Transit Analyzer (CTA). Salient results included: (1) significantly increased RBC levels of Hbm following ionophore treatment; (2) elevated Hbm with increasing lysing medium calcium concentration (0-5 mM); (3) decreased deformability of both intact RBC and ghosts with increasing Hbm and significant (P less than 0.02 or better) linear relationships between Hbm and RBC or ghost transit times; and (4) an increased sensitivity to ionophore treatment/membrane attached hemoglobin for the higher percentiles of the CTA transit time distribution (i.e., for more rigid subpopulations). Our results thus indicate that calcium-induced interaction of hemoglobin with the RBC membrane produces cellular rheological changes; in addition, they demonstrate the usefulness of the CTA system in measuring both average RBC rheologic behavior and the distribution of cellular rheologic properties within an erythrocyte population.

Pulse shape analysis of RBC micropore flow via new software for the cell transit analyser (CTA).

The Cell Transit Analyser (CTA) provides a means to rapidly measure the deformability of large numbers of individual cells. It combines many of the advantages of micropipette studies with the simplicity and speed of filtrometry methods by measuring the duration of each resistive pulse generated as a cell passes through one of 30 identical micropores in a membrane. However, in our opinion, the potential of the system is limited by the microcomputer and software supplied for data analysis. We have therefore written new software for a more-powerful microcomputer to examine the shape of each resistive pulse rather than just the duration. Seven new parameters are derived, which provide additional information regarding the passage of cells through the pores. In particular, the contribution of the entry and exit phases of the cell transit are evident in the rise time and fall time of the pulses. The software is user-friendly and allows the analysis of each pulse to be reviewed, which aids understanding of the system and helps to avoid errors in interpreting the data.

Individual red blood cell (RBC) transit analysis through sized micropores of oligopore filters : Application to RBC subpopulations

Abstract To study the transit times of individual red blood cells passing through cylindrical micropores and in order to evaluate sub-populations of cells with respect to their deformability, we have developed a new system, the Cell Transit Analyser (CTA). This computer assisted device provides the distribution of transit times τ for 5000 cells in about 1 minute. Experiments with red cells were designed to evaluate the flow behavior of both normal and abnormal cells. A statistical analysis of the results indicates a significant difference between control and rigid cells.

Increased erythrocyte rigidity in chronic alcoholics without cirrhosis: Deformability improvement of erythrocyte sub-populations after alcohol withdrawal

Red blood cell (RBC) deformability in chronic alcoholism has been studied, using the Cell Transit Analyzer (CTA) which provides transit times of 5000 individual RBC passing via cylindrical micropores of 5.0 pm in diameter and 15 pm in length. Blood from chronic alcoholic patients without cirrhosis was studied before and after 8 days alcohol withdrawal. The results indicate: 1) the mean transit time of the RBC population from alcoholic patients is greater than those of the control population, 2) these changes affect the entire cell population and not only a sub-population, 3) alcohol withdrawal for 8 days improved the deformability of the most rigid cells of alcoholic patients (i.e., a slight but significant decrease of the highest percentile values of the transit time distribution).

A new micropore filtration approach to the analysis of white cell rheology

Transit times of individual polymorphonuclear leukocytes (PMN) through cylindrical micropores with a diameter of 8 microns and a length of 19 microns were measured at pressure gradients of 3 to 10 cm H2O using the Cell Transit Analyzer (CTA); this system employs a conductometric principle and provides a frequency histogram of transit times for greater than 1000 cells within 3 to 4 minutes. Salient results included: 1) linear pressure-flow relations (r greater than or equal to 0.99); 2) mean transit times for PMN on the order of 10-20 milliseconds at a pressure gradient of 4 cm H2O and thus transit times at least 10-fold longer than for RBC under the same conditions; 3) broad distributions with marked skewness toward longer transit times, indicating a wide range of rheologic properties; 4) increased transit times in either hypotonic or hypertonic media; 5) marked increases of transit times following activation with FMLP; 6) considerable differences between the volume distribution and the transit time distribution of PMN. These results suggest the basic usefulness of the CTA for the analysis of rheologic properties of white blood cells in both normal and pathologic states.