Rui Lima

Confocal micro-PIV measurements of three-dimensional profiles of cell suspension flow in a square microchannel

A detailed measurement of the blood flow velocity profile in microchannels in vitro is fundamental to better understand the biomechanics of microcirculation. Therefore it is very important to determine the influence of suspended blood cells on the flow behaviour with high accuracy and spatial resolution. We measured the flow of blood cells suspended in a physiological fluid within a square microchannel using a confocal particle image velocimetry (PIV) system and compared it to pure water. This emerging technology combines a conventional PIV system with a spinning confocal microscope and has the ability to obtain high-resolution images and three-dimensional (3D) optical section velocity measurements. The good agreement obtained between the measured and estimated results suggests that macroscale flow theory can be used to predict the flow behaviour of a homogeneous fluid within a 100 µm square microchannel. Our results also demonstrated the potential of the confocal system for generating 3D profiles and consequently obtaining detailed information on microscale effects in microchannels using both homogeneous and non-homogeneous fluids, such as a suspension of blood cells. Furthermore, the results obtained from our confocal micro-PIV system show the ability of this system to measure velocities up to 0.52 mm s−1 in a blood cell suspension fluid.

Measurement of individual red blood cell motions under high hematocrit conditions using a confocal micro-PTV system.

Developments in optical experimental techniques have helped in elucidating how blood flows through microvessels. Although initial developments were encouraging, studies on the flow properties of blood in microcirculation have been limited by several technical factors, such as poor spatial resolution and difficulty obtaining quantitative detailed measurements at such small scales. Recent advances in computing, microscopy, and digital image processing techniques have made it possible to combine a particle tracking velocimetry (PTV) system with a confocal microscope. We document the development of a confocal micro-PTV measurement system for capturing the dynamic flow behavior of red blood cells (RBCs) in concentrated suspensions. Measurements were performed at several depths through 100-μm glass capillaries. The confocal micro-PTV system was able to detect both translational and rotational motions of individual RBCs flowing in concentrated suspensions. Our results provide evidence that RBCs in dilute suspensions (3% hematocrit) tended to follow approximately linear trajectories, whereas RBCs in concentrated suspensions (20% hematocrit) exhibited transversal displacements of about 2% from the original path. Direct and quantitative measurements indicated that the plasma layer appeared to enhance the fluctuations in RBC trajectories owing to decreased obstruction in transversal movements caused by other RBCs. Using optical sectioning and subsequent image contrast and resolution enhancement, the system provides previously unobtainable information on the motion of RBCs, including the trajectories of two or more RBCs interacting in the same focal plane and RBC dispersion coefficients in different focal planes.

Radial dispersion of red blood cells in blood flowing through glass capillaries: the role of hematocrit and geometry

The flow properties of blood in the microcirculation depend strongly on the hematocrit (Hct), microvessel geometry, and cell properties. Previous in vitro studies have measured the radial displacement of red blood cells (RBCs) at concentrated suspensions using conventional microscopes. However, to measure the RBCs motion they used transparent suspensions of ghost red cells, which may have different physical properties than normal RBCs. The present study introduces a new approach (confocal micro-PTV) to measure the motion of labeled RBCs flowing in concentrated suspensions of normal RBCs. The ability of confocal systems to obtain thin in-focus planes allowed us to measure the radial position of individual RBCs accurately and to consequently measure the interaction between multiple labeled RBCs. All the measurements were performed in the center plane of both 50 and 100 microm glass capillaries at Reynolds numbers (Re) from 0.003 to 0.005 using Hcts from 2% to 35%. To quantify the motion and interaction of multiple RBCs, we used the RBC radial dispersion (D(yy)). Our results clearly demonstrate that D(yy) strongly depends on the Hct. The RBCs exhibited higher D(yy) at radial positions between 0.4 and 0.8R and lower D(yy) at locations adjacent to the wall (0.8-1R) and around the middle of the capillary (0-0.2R). The present work also demonstrates that D(yy) tends to decrease with a decrease in the diameter. The information provided by this study not only complements previous investigations on microhemorheology of both dilute and concentrated suspensions of RBCs, but also shows the influence of both Hct and geometry on the radial dispersion of RBCs. This information is important for a better understanding of blood mass transport mechanisms under both physiological and pathological conditions.

Microscale Flow Dynamics of Red Blood Cells in Microchannels: An Experimental and Numerical Analysis

The blood flow dynamics in microcirculation depends strongly on the motion, deformation and interaction of red blood cells (RBCs) within the microvessel. We present confocal micro-PTV measurements on the motion of individual RBCs through a circular polydimethysiloxane (PDMS) microchannel. The RBC radial displacement and dispersion calculated from these measurements show that the RBC paths are strongly dependent on the both Hct and plasma layer. In order to obtain more detailed information of the non-Newtonian property of blood a novel computational scheme is also described. The simulated flow dynamics were in good agreement with the Casson flow model and in vivo observations. In the near future by comparing both results we hope to clarify a variety of complex phenomena occurring at the microscale level.

Velocity measurements of blood flow in a rectangular PDMS microchannel assessed by confocal micro-PIV system

This paper examines the ability to measure the velocity of both physiological saline (PS) and in vitro blood in a rectangular polydimethysiloxane (PDMS) microchannel by means of the confocal micro-PIV system. The PDMS microchannel, was fabricated by conventional soft lithography, had a microchannel near to a perfect rectangular shape (300µm wide, 45µm deep) and was optically transparent, which is suitable to measure both PS and in vitro blood using the confocal system. By using this latter combination, the measurements of trace particles seeded in the flow were performed for both fluids at a constant flow rate (Re=0.021). Generally, all the velocity profiles were found to be markedly blunt in the central region mainly due to the low aspect ratio (h/w=0.15) of the rectangular microchannel. Predictions by a theoretical model for the rectangular microchannel have showed fairly good correspondence with the experimental micro-PIV results for the PS fluid. Conversely, for the in vitro blood with 20% haematocrit, small fluctuations were found on velocity profiles.

Confocal Micro-PIV/PTV Measurements of the Blood Flow in Micro-channels

The development of optical experimental techniques has contributed to obtaining explanations of the behaviour of blood flowing in micro-channels. Although past results have been valuable, detailed studies on the flow properties of in vitro blood in micro-channels have been limited by several technical factors such as poor spatial resolution and difficulty in obtaining quantitative detailed measurements at such small scales. In recent years, due to advances in computers, optics, and digital image processing techniques, it has become possible to combine both particle image velocimetry (PIV) and particle tracking velocimetry (PTV) methods with confocal microscopes. As a result, this combination has greatly increased the resolution of conventional micro-PIV/PTV systems and consequently provided additional detailed description on the motion of blood cells not obtainable by traditional methods. In this chapter the most relevant theoretical and technical issues related to both conventional and confocal micro-PIV/PTV methods are discussed. Additionally, the most recent studies on the blood flow behaviour in micro-channels obtained by our confocal micro-PIV/PTV system are also reviewed.

105 DISPERSION OF RED BLOOD CELLS IN MICROCHANNELS : A CONFOCAL MICRO-PTV ASSESSMENT

1) Dept. Bioeng. & Robot, G. S. Eng., Tohoku Univ., 6-6-01 Aoba, 980-8579 Sendai, Japan. 2) Dept. Mech. Tech., ESTiG, Braganca Polyt., C. Sta. Apolonia, 5301-857 Brag., Portugal. 3) Div. Surgical Oncology, Tohoku Univ., 2-1 Seiryo-machi, 980-8575 Sendai, Japan. 4) Dept. Mech. Sci. and Bioeng., G. S. Eng., Osaka Univ., 560-8531 Osaka, Japan. 5) New Industry Hatchery Centre, Tohoku Univ., 6-6-01 Aoba, 980-857, Sendai, Japan.

P-01 OBSERVATION OF THE BLOOD FLOW IN MICROCHANNEL WITH STENOSIS BY CONFOCAL-MICRO-PIV

Introduction Mass transport in human cardiovascular system takes place mainly in microcirculation, so it is important to understand blood !low in microvessels. In such small microchannels, behaviors of red blood cells(RBCs) strongly affect the now lie! d. Recently many researcher developed various systems to visualize blood !low in microchannels. But it is very difficult to visualize the centcr plane or the blood flow when the concentration of RBCs(HCT) is over I 0%. In our study, we used confocal-micro-PIV and PTV systems to overcome this problem, and investigate blood flow through stenosis.

Measurement of Erythrocyte Motions in Microchannels by Using a Confocal Micro-PTV System

Detailed knowledge on the motion of individual red blood cells (RBCs) flowing in microchannels is essential to provide a better understanding on the blood rheological properties and disorders in microvessels. Several studies on both individual and concentrated RBCs have already been performed in the past [1, 2]. However, all studies used conventional microscopes and also ghost cells to obtain visible trace RBCs through the microchannel. Recently, considerable progress in the development of confocal microscopy and consequent advantages of this microscope over the conventional microscopes have led to a new technique known as confocal micro-PIV [3, 4]. This technique combines the conventional PIV system with a spinning disk confocal microscope (SDCM). Due to its outstanding spatial filtering technique together with the multiple point light illumination system, this kind of microscope has the ability to obtain in-focus images with optical thickness less than 1 μm, a task extremely difficult to be achieved by using a conventional microscope.Copyright

VELOCITY FIELDS OF BLOOD FLOW IN MICROCHANNELS USING A CONFOCAL MICRO-PIV SYSTEM

This study was supported in part by the following grants: 21st Century COE Program for Future Medical Engineering based on Bio-nanotechnology, International Doctoral Program in Engineering from the Ministry of Education, Culture, Sports, Science and Technology of Japan (MEXT), “Revolutionary Simulation Software (RSS21)” next-generation IT program of MEXT; Grants-in-Aid for Scientific Research from MEXT and JSPS Scientific Research in Priority Areas (768) “Biomechanics at Micro- and Nanoscale Levels,” Scientific nResearch (A) No.16200031 “Mechanism of the formation, destruction, and movement of thrombi responsible for ischemia of vital organs.” The authors also thank all members of Esashi, Ono and Tanaka Lab. for their assistance in fabricating the PDMS microchannel.