How can rbc volume be measured

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Abstract IN both clinical practice and, experimental research, isotope dilution methods 1—7 are being used routinely for the determination of total blood volume. Access through your institution. Buy or subscribe. Rent or Buy article Get time limited or full article access on ReadCube. References 1 Larchan, A. Article Google Scholar 2 Andersen, S. Google Scholar 3 Agarwal, S.

Google Scholar 10 Bernhard, W. Google Scholar 11 Fudenberg, H. Article Google Scholar 15 Gurney et al. A Topogram of normal RBCs. B Detailed texture of the RBC membrane surface. C Indentation depth measurement.

D Different force-versus-indentation depth curves of RBCs in various conditions: a. G6PD deficiency; and n. Reproduced, with permission, from Kamruzzahan, Kienberger et al. The displacement of the stage required for the same deflection of the tip is different between solid- and soft-materials, from which applied forces can be calibrated.

The Poisson ratio is 0. RBCs from hereditary spherocytosis, thalassemia Dulinska, Targosz et al. Light refraction at a sample induces linear momentum change, resulting into trapping forces Fig. High numerical aperture NA objective lens is used to generate a tightly focused optical trap, and its trapping force is governed by the refractive indices of sample and surrounding medium, laser power, and sample size; optical force to trap particles much smaller than laser wavelength can be described by Rayleigh scattering theory, while trapping samples much larger than laser wavelength belongs to Mie scattering regime Ashkin, Dziedzic et al.

Principles of optical tweezers. A Laser beam with gradual intensity transfers linear momentum to a microsphere to escape from the beam center. B Focused Gaussian beam exerts trapping force.

Modified, with permission, from Svoboda and Block, ; Henon, Lenormand et al. Since optical tweezers can apply forces at the pN level, it has been employed for measuring the deformability of RBCs.

Measurements of the mechanical properties of RBCs with optical tweezers can be done either by applying optical force to microspheres attached to RBCs Henon, Lenormand et al. The change in the projected diameter of the RBC in response of optical force is converted to shear modulus of the RBC using mathematical membrane models. Optical stretcher, a variant of optical tweezers, uses two diverging laser beams from opposite directions Guck, Ananthakrishnan et al.

Linear momentum changes by two laser beams apply stretching force to the RBC along the optical axis, and the RBC deformations under varying optical force are measured from which mechnical properties are retrieved. Optical tweezers can also be used for detecting membrane fluctuation dynamics of RBCs by imposing a deformation Yoon, Kotar et al.

Magnetic twisting cytometry MTC applies both static and oscillating magnetic field to ferromagnetic microbeads attached to the surface of cell membrane Wang, Butler et al. Depending on the applied magnetic field, the microbeads exhibit both translational and rotational motion, which applies torques to the cell membrane.

By varying oscillating frequency 0. Reproduced, with permission, from Puig-de-Morales-Marinkovic, Turner et al. Quantitaitve phase imaging technqiues measure the electric field, i. Employing the principle of laser interference, electric field information of target sample is modulated onto intereferograms recorded by a CCD camera. By using appropriate field retrieval algorithms, the field information can be retrieved from the measured holograms Debnath and Park, Typical interferogram and quantitative phase image of a RBC are shown in Fig.

Quantitative phase imaging techniques can measuring dynamic membrane fluctuations of RBCs Popescu, Ikeda et al. Dynamic membrane fluctuation, consisting of submicron displacement of the membrane, has a strong correlation with deformability of RBCs and can be altered by biochemical changes in protein level Waugh and Evans, Quantitative phase imaging. A Schematic of the principle of quantitative phase imaging.

Reproduced, with permission, from Park, Best et al. Diffraction phase microscopy DPM , a highly stable technique for quantitative phase imaging, has been widely used for investigating the deformability of RBCs. Employing common-path laser interferometry, DPM provides full-field quantitative phase imaging with unprecedented stability Park, Popescu et al. DPM measured spatiotemporal coherency in dynamic membrane fluctuations Popescu, Park et al.

Recently, integrated with a mathematical model, DPM provide the mechanical properties of individual RBCs from membrane fluctuations: shear modulus, bending modulus, area expansion modulus, and cytoplasmic viscosity Park, Best et al. Employing spectroscopic quantitative phase imaging, cytoplasmic Hb concentration that is tightly related to the cytoplasmic viscosity, can also be simultaneously quantified Park, Yamauchi et al.

In addition, polarization-sensitive quantitative phase microscopy will be potentially used for the study of sickle cell disease and its implications to RBC deformability Kim, Jeong et al.

Although dynamic light scattering have been extensively used in combination with ektacytometry, it provides averaged signals from many RBCs. Thus it is difficult to access the deformability of individual RBCs. Fourier transform light scattering FTLS provides both static and dynamic light scattering signal from individual cells. Light field, measured by quantitative phase microscopy or digital holographic microscopy, contains both amplitude and phase information, and thus far-field light scattering pattterns can be directly calculated by numerically propagating the measured field — technically applying Fourier transformation Ding, Wang et al.

FTLS technique can provide both morphological and rheological information about individual biological cells. Due to its capability of measuring light scattering signals from individual cells with high signal-to-noise ratio, FTLS has been employed to study several pathophysiological effects to the deformabiltiy of RBCs, including malaria infection Park, Diez-Silva et al. Blood viscometer measures the viscosity of blood over a wide range of shear rates. Blood viscometer controls either shear stress or shear rate of blood using rational objects.

Stress-controlled blood viscometer applys a constant torque which corresponds to constant rotational speed in a well-designed rotational rheometer. In a rate-controlled system, applied torque is controlled by a stress-sensing device so that a constant rotational speed is achieved. Viscometers can be classified by the cylinder shape: a concentric cylinder, a cone plate, and a parallel plate viscometer Fig.

Cylinder-type viscometer uses two concentric cylinders: a rotational inner cup and a stationary outer cylinder. Time-independent shear rate can be precisely measured by concentric cylinder viscometer Nguyen and Boger, Parallel plate viscometer is a simplified version of the cone and plate viscometer and has a advantage of flexible space between two parallel plates. The viscous fluid can confined and rotated in narrow space between two circular parallel plates Gent, Schematic diagrams of typical viscometers.

Ektacytometer employes a laser diffraction technique with blood viscometer in order to measure RBC deformabiltiy. Conventional blood viscometer applys controlled shear stress to the RBCs in the blood viscometer, and deformability of RBCs can be measured from laser diffraction pattern. Ektacytometer consists of a concentric rotational outer cup and a stationaly inner cylinder; outer cup produces varying shear stress field on blood Fig.

Through the measurement of diffraction patterns of the laser passing through the blood, RBC deformability can be obtained. The RBC deformation is quantitatively calculated from the scattered laser beam intensity pattern. Under a certain shear rate, isointensity curves in the intensity pattern of the scattered beam will show elliptical shapes, which represent elliptically deformed RBC population Bessis, Mohandas et al. DI values are measured at different angular velocities and thus different shear rate of the outer cyliner in the ektacytometer.

Ektacytometer is a simple and effective technique to measure the deformability of RBC population, and it has been widely used for the study pathophysiology of RBCs. Abnormal deformability in RBCs from patients with hereditary pyropoikilocytosis, hereditary spherocytosis, and Hb CC disease were studies by ektacytometer Mohandas, Clark et al.

Microfludic device reduced space, labor, and measurement time on numerous experiments, and also enabled precise control and manipulation of the small volume of samples.

Microfludic device has been used to study the deformabiltiy of RBCs. Microfluidic channel with a few micrometer diameter mimics micro-capillary structure in blood circulation system. Microfludic device was used to induce large deformation of RBCs and its mechanical behavior was studied Fig. Reproduced, with permission, from Li, Lykotrafitis et al.

For the study of sickle cell disease, microfluidic device has been used to measure the resistance change rate of blood flow under the sudden change of oxygen concentration Wang, Ding et al.

Recently, microfludic channels with obstracles have measured the deformabiltiy of malaria infected RBCs in high throughput Bow, Pivkin et al. By applying a negative pressure, whole blood is subject to pass through holes in the membrane filter. The deformability of RBCs can affect the speed of flow. Since the filteration test requires for a relatively simple instrument and provides clinically relevant results with high reproducibility, it has been widely used in various studies related to RBC deformability, including the effects of diabetes Juhan, Buonocore et al.

This essential deformability is in turn affected by various physiological and pathological cues. Temperature plays important roles in RBC deformabilty.

The elastic properties of RBC membrane were investigated as function of temperature using the micropipette aspiration technique Waugh and Evans, Due to the structual transitions of proteins occuring at certain critical tempertures, RBC deformabiltiy exhibits complex behaviors. Body temperature or febril temperature are particularly important in various pathophysiology of RBCs.

Membrane fluctuation measurements using DPM revealed that the shear modulus of Pf-RBCs significantly increases as temperature increases from body temperature to febrile temperature whereas healthy RBCs do not show noticible changes Park, Diez-Silva et al. RBCs exhibit diverse morphological features depending on pathophysiological conditions Diez-Silva, Dao et al. A healthy human RBC shows a smooth and biconcave disc shape discocyte. However, atypical shapes of RBCs can be found under abnormal pathophysiological conditions, including acanthocyte, stomatocyte, schizocyte, and tear drop cells Kenneth, One of the hypotheses describing RBC morphology is the bilayer-couple hypothesis Sheetz and Singer, ; small changes in the relaxed area difference between two layers of phospholipids.

Later, this model can be used for explaning stomatocyte—discocyte—echinocyte morphological transitions Lim HW, Wortis et al.

Increased deformability of RBCs in abnormal shapes has been reported with various experimental methods. The shear moduli values show bimodal distributions Fig. The aged RBCs were found by ektacytometry to have decreases shear modulus mainly because of decreased surface area and increased cytoplasmic viscosity.

Different osmolalities of extracellular medium can bring significant changes in RBC shape and thus deformability. In hypotonic medium, RBCs are swollen due to water intake. In the hypertonic case, RBCs lose its volumes, which result in significant cell shrinkage. Although the total amount of Hb molecules in RBCs, or the mean corpuscular Hb MCH , does not significantly change at different osmolality, Hb concentration can be considerably changed due to water influx and efflux.

RBCs exhibit the maximum deformability at physiological condition; under either hypertonic or hypotonic condition, the deformability of RBCs decreases Mohandas, Clark et al. The following steps describe what happens:. The RBC indices can help your doctor determine the cause if your doctor thinks you have anemia. Your doctor will see if your MCV is low, normal, or high to help determine what is causing your anemia.

The MCV is higher than normal when red blood cells are larger than normal. This is called macrocytic anemia. The MCV will be lower than normal when red blood cells are too small.

This condition is called microcytic anemia. If you have a normal MCV, it means that your red blood cells are normal in size. You can have a normal MCV and still be anemic if there are too few red blood cells or if other RBC indices are abnormal. This is called normocytic anemia. Normocytic anemia occurs when the red blood cells are normal in size and hemoglobin content, but there are too few of them. This can be caused by:. If you have a high MCHC, this means that the relative hemoglobin concentration per red blood cell is high.

MCHC can be elevated in diseases such as:. If you have a low MCHC, it means that the relative hemoglobin concentration per red blood cell is low. The red blood cells will take on a lighter color when viewed under the microscope. Individuals with anemia and a corresponding low MCHC are said to be hypochromic. Your doctor may also perform other tests to make a diagnosis. Treatment for any anemia depends on the underlying cause. For example, if your anemia is caused by iron deficiency, your doctor may advise you to take iron supplements or change your diet to include more foods that are rich in iron.

The test is usually part of a complete blood count…. Understand anisocytosis, its relation to anemia, and how it is typically diagnosed and treated. While low Hgb generally indicates anemia, high Hgb may be caused by a….