A microfluidic device for partial cell separation and deformability assessment

Blood flow in microcirculation shows several interesting phenomena that can be used to develop microfluidic devices for blood separation and analysis in continuous flow. In this study we present a novel continuous microfluidic device for partial extraction of red blood cells (RBCs) and subsequent measurement of RBC deformability. For this purpose, we use polydimethylsiloxane (PDMS) microchannels having different constrictions (25%, 50% and 75%) to investigate their effect on the cell-free layer (CFL) thickness and separation efficiency. By using a combination of image analysis techniques we are able to automatically measure the CFL width before and after an artificial constriction. The results suggest that the CFL width increases with enhancement of the constriction and contributes to partial cell separation. The subsequent measurements of RBCs deformation index reveal that the degree of deformation depends on the constriction geometries and hematocrit after the cell separation module. The proposed microfluidic device can be easily transformed into a simple, inexpensive and convenient clinical tool able to perform both RBC separation and deformability analysis in one single device. This would eliminate the need for external sample handling and thus reducing associated labor costs and potential human errors

Analysis of the cell-free layer in a circular microchannels: trajectories of labeled red bllod cells

In this experimental work, we measure the trajectories of the cell-free layer (CFL) by tracking labeled red blood cells (RBCs) flowing around the boundary of the RBCs core. The circular glass microchannels studied are 100 um in diameter. The images are captured using confocal system and are post-processed using Image J and MATLAB. The results suggest that the trajectories follows a polynomial function

Erythrocyte deformability in a hyperbolic microchannel

Serious diseases such as diabetes and malaria are deeply related to microcirculation disorders caused by abnormal erythrocytes deformability [1]. Therefore, various studies on red blood cells (RBCs) deformation have been carried out but those studies rather focus on shear ow or extensional ow [2-6]. In this study, a relatively low aspect ratio (AR) hyperbolic microchannel was used in order to create an extensional ow combined with a shear ow, where AR= h w , h means the depth and w means the width of the microchannel. The objective of the study is to investigate the degree of RBC deformation throughout the microchannel at the centerline (y = 0). The blood samples were RBCs diluted with the Hank's Balanced Salt Solution (HBSS) with the hematocrit (Hct) level set to be 2%. A polydimethylsiloxane (PDMS) microchannel, having a hyperbolic contraction region followed by an abrupt expansion shape, was fabricated by a soft-lithography technique. The dimensions of the microchannel: width of the inlet (w), length of contraction region (l), width of the exit of contraction (wc) and depth (h) were 400 m, 580 m, 20 m and 14 m, respectively. Hence, the AR was 0:035. The experimental equipment consisted of an inverted microscope (Diaphot 300, Nikon), a high-speed camera (FASTCAM SA3, Photron) connected to a computer and a 1mL syringe (TERUMO R SYRING) controlled by a syringe pump (PHD ULTRA). The manufactured PDMS microchannel was placed on the stage of the microscope where the ow rate Q of the working uids was kept constant at 0:5 L=min. The owing RBCs were recorded by a high speed camera at a frame rate of 7500 frames/s and analyzed. The DI was obtained by DI= a b a + b where a refers to the major axis and b refers to the minor axis of the ellipse best tted to RBCs. High DI values in the contraction region were observed. It is possible to say that the RBCs highly elongate with both extensional and shear dominated ows achieved by a hyperbolic microchannel with a relatively low aspect ratio AR

Extensional flow-based microfluidic device: deformability assessment of red blood cells in contact with tumor cells

Red blood cell (RBC) deformability has become one of the important factors to assess blood and cardiovascular diseases. The interest on blood studies have promoted a development of various microfluidic devices that treat and analyse blood cells. Recent years, besides the RBC deformability assessment, these devices are often applied to cancer cell detection and isolation from the whole blood. The devices for cancer cell isolation rely mainly on size and deformability of the cells. However, the examination of deformability of the RBCs mixed with cancer cells is lacking. This study aims at determining the deformation index (DI) of the RBCs in contact with cancer cells using a hyperbolic microchannel which generates a strong extensional flow. The DIs of human healthy RBCs and human RBCs in contact with a tumor cell line (HCT-15, colon carcinoma) were compared by analyzing the flowing RBCs images captured by a high speed camera. The results reveal that the RBCs that were in contact with HCT-15 cells have lower deformability than the normal RBCs.The authors acknowledge the financial support provided by: Student Mobility Placements with the program Lifelong Learning (Erasmus Program), 2007 Global COE Program “Global Nano-Biomedical Engineering Education and Research Network”, Japan. Grant-in-Aid for Science and Technology (PTDC/SAU-BEB/105650/2008, PTDC/EME-MFE/099109/2008 and PTDC/SAU-ENB/116929/2010) from the Science and Technology Foundation (FCT) and COMPETE, Portugal. The authors are also very grateful to Professor Mónica S.N. Oliveira (Strathclyde University), Professor Geyong M. Kim (University of Navarra) and Professor Sergio Arana (University of Navarra) for their discussion and suggestions to this research work

Visualization and measurement of red blood cells flowing in microfluidic devices

Several experimental techniques were performed in the past years using in vitro environments, in an attempt to not only understand the blood flow behaviour in microcirculation but also develop microfluidic devices as an alternative clinical methodology to detect blood diseases. Hence, the visualization and measurement of red blood cells (RBCs) flowing in a microfluidic device are important to provide not only essential information about hydrodynamic characteristics of the blood but also vital information to diagnose the initial symptoms of diseases during clinical investigations. For instance, RBC rigidity has been correlated with myocardial infarction, diabetes mellitus, hypertension, and also other haematological disorders and diseases that affect RBC deformation more directly, such as, hereditary spherocytosis, sickle cell anaemia and malaria. Regarding a better understanding of the RBCs deformation and motion, we present in this paper a compilation of studies made in our research group, using several microfluidic devices with different microchannel geometries and fabrication techniques (i.e., soft-lithography, xurography and hybrids) that focus in the shear and extensional flow behaviour, either in healthy or chemically stiffed RBCs.The authors acknowledge the financial support provided by 2007 Global COE Program “Global Nano-Biomedical Engineering Education and Research Network”, Japan and grant-in-Aid for Science and Technology, PTDC/SAU-BEB/105650/2008, PTDC/EME-MFE/099109/2008, PTDC/SAUENB/116929/2010, EXPL/EMS-SIS/2215/2013, scholarship SFRH/BD/91192/2012 and scholarship SFRH/BD/89077/2012 from FCT (Science and Technology Foundation), COMPETE, QREN and European Union (FEDER). The authors are also very grateful to Dr. Mónica Oliveira (Strathclyde University), Professor Takuji Ishikawa and Professor Takami Yamaguchi (Tohoku University) for their suggestions and support to this research work

Visualization of the cell-free layer (CFL) in a PDMS microchannel with a micro-stenosis

Red blood cells (RBCs) have a tendency to undergo axial migration due to the parabolic velocity profile which results in a high shear stress around wall that forces the RBC to move towards the center induced by the tank treading motion of the RBC membrane. As a result there is a formation of cell-free layer (CFL) with extremely low concentration of cells. Based on this phenomenon several works have proposed microfluidic designs to separate the suspending physiological fluid from whole in vitro blood. However, most of these studies have the aim of the complete extraction of cells from plasma which is not the case of the present study. The biomedical device that is present in this work aims to obtain a CFL with a low enough RBC propose a combination of image analysis techniques able to measure automatically the CFL thickness before and after micro-stenosis is used

Separation and deformation of red blood cells in PDMS microchannels

Over the years, several experimental techniques were performed in in vitro environments, in an attempt to understand the flow behaviour of blood in microcirculation. Several of these studies were performed in glass capillaries, and have produced significant results with respect to rheological properties of blood [1, 2]. Another way to perform in vitro blood studies is to use microchannels fabricated by soft- lithography [3, 4] and xurography [5]. With these techniques several studies have focused in the formation of the cell-free layer (CFL) that is caused by the tendency of red blood cells (RBCs) to migrate toward the centre of the microchannel, in that the physical reason is known as the Fahraeus Lindqvist. The presence of this CFL at the regions adjacent to the wall is affected by the geometry of the microchannel [4] and the physiological conditions of the working fluid, such as the hematocrit (Hct) [6], and the RBC deformability [7]. The formation of CFL can be used for separation of diseased cells from healthy blood cells [8]. The aim of this paper is to show briefly the importance of the microfluidic devices to study several physiological phenomena that happens in vivo environments with special focus on the CFL behaviour and RBC deformability

Imaginando o Japão a partir do Extremo Oeste: representações sociais do Japão e dos japoneses entre estudantes portugueses

Tese de doutoramento em Estudos CulturaisIn the educational and academic field related to Japanese language and culture in Portugal, there is a need to understand how to approach “Other/Oriental” bearing in mind socio-cultural phenomena and human perception. The principal objective of this study is to analyze how Japan, the Japanese and Japanese culture are socially represented among Portuguese students, more specifically in Japanese as a Foreign Language (JFL) context. Two empirical studies were conducted after articulating the several concepts developed in different disciplines such as Cultural Studies, Japanese Studies, Media and Communication, and Social Psychology. For the first study, we have conducted a questionnaire survey (N=545) to JFL and non-JFL students and have examined Japan’s representation and social stereotypes about the Japanese. We were thus able to identify the centrality of popular culture in social representations of Japan. Through a factor analysis, five factors related to the Japanese characteristics were identified, “Politeness”, “Diligence”, “Shyness”, Conservativeness” and “Innovativeness”. Then, seven focus groups were organized and seven themes came up by examining the students’ discussion about a transnational cultural object, anime (Japanese animation). The results suggest that the students represent old and new aspects of Japan, which may correspond to Orientalized and Techno-Orientalized perceptions of Japan. It was also observed that there are stereotypes of the Japanese that are cross-culturally shared by Portuguese students, such as diligence, which may underline the persistence of some images that were probably constructed in the period of Japan’s rapid economic growth in the 60s and 70s. However, as the JFL group has indicated more variability in the traits ascription than the non-JFL group, familiarization with Japanese culture through human communication may contribute to reconstruct the representations of Japan. These findings lead us to acknowledge the importance of intercultural dialogues. Considering today’s easy and rapid transnational circulation of information, it is essential for the researchers and educators who engage in mediating knowledge production and intercultural teaching to be aware of the continuity and discontinuity of cultural and social norms and values.No domínio pedagógico e académico relacionado com a língua e cultura japonesa em Portugal, é necessário compreender como abordar o “Outro/Oriental” tendo em conta os fenómenos socioculturais e a percepção humana. O objetivo principal do presente estudo é analisar como o Japão, os japoneses e a cultura japonesa são socialmente representados entre estudantes portugueses, em particular no contexto do Japonês como Língua Estrangeira (JLE). Tendo em conta a articulação de vários conceitos desenvolvidos em diferentes áreas de estudo, tais como os Estudos Culturais, Estudos Japoneses, Media e Comunicação e Psicologia Social, foram realizados dois estudos empíricos. No primeiro estudo, foi realizado um questionário (N = 545) dirigido a alunos JLE e não JLE e foram examinadas as representações do Japão e os estereótipos sociais dos japoneses. Neste estudo, foi identificada a centralidade da cultura popular nas representações sociais do Japão. Por intermédio de uma análise fatorial, foram identificados cinco fatores para as características dos japoneses, a saber, “Polidez”, “Diligência”, “Timidez”, “Conservadorismo” e “Inovação”. Numa segunda fase, foram organizados sete grupos focais e foram identificados sete temas após análise da discussão dos alunos sobre um objeto cultural transnacional, o anime (animação japonesa). Os resultados deste estudo sugerem que os alunos representam aspetos antigos e novos do Japão, o que corresponde a imagens Orientalizadas e Tecno-Orientalizadas do Japão. Também se verificou que existem estereótipos dos japoneses transculturalmente partilhados entre os estudantes portugueses, como por exemplo a diligência, o que pode sublinhar a persistência de algumas das imagens que provavelmente foram construídas no período do rápido crescimento económico do Japão nos anos 60 e 70. No entanto, como o grupo JLE mostrou mais variabilidade na atribuição de características do que o grupo não JLE, a familiarização com a cultura japonesa pela comunicação humana pode ser um fator relevante para a reconstrução das representações do Japão. Estes resultados sugerem a importância de reconhecer os diálogos interculturais. Tendo em conta a fácil e rápida circulação transnacional de informações hoje em dia, é essencial que os investigadores e educadores envolvidos na produção de conhecimento e no ensino intercultural, tenham em consideração a continuidade e descontinuidade das normas e valores culturais e sociais

Motion of rigid particles flowing in a microfluidic device with a pronounced stenosis: trajectories and deformation index

Studies of in vitro blood flow behaviour in stenosis are essential not only as an attempt to understand this phenomenon, but also to develop microfluidic devices, as an alternative clinical methodology to detect blood diseases. Regarding a better understanding of the role that rigid Red Blood Cells (rRBCs) play in the blood flow behaviour, we have studied the trajectories and deformation of both, artificial rigid Red Blood Cells (arRBCs) and healthy ovine Red Blood Cells (RBCs), when subjected to a pronounced microstenosis. By using a high-speed video microscopy system, the present study was performed with different flow rates (1, 10 and 20 µl/min). The results have shown that healthy RBCs and arRBCs present deviations in their flow behaviour and deformation index, which might be a potential alternative clinical methodology to detect blood cells related diseases.The authors acknowledge the financial support provided by PTDC/SAU-BEB/105650/2008, PTDC/EME-MFE/099109/2008, PTDC/SAU-ENB/116929/2010, EXPL/EMS-SIS/2215/2013, scholarship SFRH/BD/91192/2012 and scholarship SFRH/BD/89077/2012 from FCT (Science and Technology Foundation), COMPETE, QREN and European Union (FEDER)

Flow of red blood cells through a microfluidic extensional device: an image analysis assessment

The present study aims to assess the deformability of red blood cells (RBCs) under extensionally dominated microfluidic flows using an image based technique. For this purpose, a microchannel having a hyperbolic shaped-contraction was used and the images were captured by a standard high-speed microscopy system. The images acquired display RBCs with various light intensity levels and image analysis was used to quantify the deformation index (DI) of the RBCs considering these light intensity differences. Additionally, the velocities of different intensity-level RBCs flowing along the centerline of the channel were measured using particle tracking velocimetry. The preliminary results at two different flow rates reveal a highly deformable nature of RBCs when submitted to strong extensional flows. It was also observed that the low intensity cells exhibit a slightly higher velocity than intermediate intensity cells, which we attribute to the cells being located in different planesWe thank Dr. Matsuki for help \Vith blood sample collection. Additionally, wc acknowledge the financiai support provided by 2007 GlobalCOE Program "Global Nano-BME Education and Research Network", Japan. We are also thankful to FCT (Portugal) and COMPETE ror financiai support through projects PTDC/SAU-BEB/I08728/2008, PTDC/SAU-BEB/105650/2008 and PTDC/EMEMFEl099I 09/2008