Label-free density difference amplification-based cell sorting

The selective cell separation is a critical step in fundamental life sciences, translational medicine, biotechnology, and energy harvesting. Conventional cell separation methods are fluorescent activated cell sorting and magnetic-activated cell sorting based on fluorescent probes and magnetic particles on cell surfaces. Label-free cell separation methods such as Raman-activated cell sorting, electro-physiologically activated cell sorting, dielectric-activated cell sorting, or inertial microfluidic cell sorting are, however, limited when separating cells of the same kind or cells with similar sizes and dielectric properties, as well as similar electrophysiological phenotypes. Here we report a label-free density difference amplification-based cell sorting (dDACS) without using any external optical, magnetic, electrical forces, or fluidic activations. The conceptual microfluidic design consists of an inlet, hydraulic jump cavity, and multiple outlets. Incoming particles experience gravity, buoyancy, and drag forces in the separation chamber. The height and distance that each particle can reach in the chamber are different and depend on its density, thus allowing for the separation of particles into multiple outlets. The separation behavior of the particles, based on the ratio of the channel heights of the inlet and chamber and Reynolds number has been systematically studied. Numerical simulation reveals that the difference between the heights of only lighter particles with densities close to that of water increases with increasing the ratio of the channel heights, while decreasing Reynolds number can amplify the difference in the heights between the particles considered irrespective of their densities.Separating specific cells from heterogeneous or homogeneous mixtures has been considered as a key step in a wide variety of applications ranging from biomedicine to energy harvesting. For example, the separation and sorting of rare circulating tumor cells (CTCs) from whole blood has gained significant importance in the potential diagnosis and treatment of metastatic cancers.^1,2 Similarly, malaria detection relies on the collection of infected red blood cells (RBCs) from whole blood.^3,4 In addition, the selective separation of lipid-rich microalgae from homogeneous mixtures of microalgae is a promising technique in biomass conversion.⁵To date, conventional cell separation can be done by labelling cells with biomolecules to induce differences in physical properties. For instance, in a fluorescence-activated cell sorter (FACS), cells to be separated are labelled with antibodies or aptamers with fluorescent molecules, and then sorted by applying an electrical potential.^6,7 Similarly, magnetic-activated cell sorter (MACS) uses magnetic.^8,9 Alternatively, label-free cell separation methods have exploited inherent differences in the physical properties (e.g., size and dielectric properties) of different kinds of cells. For example, acoustophoresis forces particles larger than a desired size to move into the center of a fluidic channel by using ultrasonic standing waves.^10–12 Inertial microfluidics takes advantage of curved fluidic channels in order to amplify the size differences between particles.^13,14 Mass-dependent separation of particles based on gravity and hydrodynamic flow was also reported.¹⁵ Particles with different dielectric properties can also be sorted by dielectrophoresis which induces the movement of polarizable particles.^16–18The disadvantage of these methods, however, is that they require external forces and labels that may cause unexpected damage to biological cells.^19–21 More importantly, most methods are limited in separating cells of the same kind or cells with similar sizes and dielectric properties.Here we designed a novel, label-free density difference amplification-based cell sorting (dDACS) that allows the separation of particles with the same size and charge by exploiting subtle differences in density without the use of external forces. Figure 1(a) illustrates the proposed microfluidic model and its underlying mechanism. The conceptual microfluidic system consists of an inlet, a separation chamber (hydraulic jump cavity), and multiple outlets. Particles entering through the inlet experience gravity (F_G), buoyancy (F_B), and drag (F_D) forces in the separation chamber. The net force acting on the particles can be described as F = F_G + F_B + F_D.(1)As particles enter the separation chamber (i.e., hydraulic jump cavity), F_D acting on the particles changes its direction along the streamline. The particles experience additional forces in the y direction due to large tangential angle (Fig. 1(b)). For lighter particles, whose densities are close to that of the surrounding water, F_D becomes comparable to F_G (i.e., in the y direction), while the net force for heavier particles is less affected by this additional contribution of F_D due to a large F_G. As a result, the height (H) and distance (D) that each particle can travel are different depending on its density. The difference in the maximum height (ΔH_max) between two particles with different density (ρ_p1 and ρ_p2) can be further approximated as ΔHmax∝(vyp0)2(vyf−vyp0), (ρp1−ρp2) ,(2)where v_yp0 and v_yf represent the velocity of particle and fluid along the y direction at the entrance of hydraulic jump cavity, respectively.Open in a separate windowFIG. 1.Schematic illustration of label-free density difference amplification-based cell sorting (dDACS), which exploits differences in the densities (ρ₁ > ρ₂) of particles with similar diameters (d) and charge. (a) The conceptual microfluidic design consists of an inlet, a separation chamber (hydraulic jump cavity), and multiple outlets. Incoming particles experience gravity (F_G), buoyancy (F_B), and drag (F_D) forces in the separation chamber, and depending on their densities, the height (H) and distance (D) that each particle is able to reach will be different, allowing the particles to be separated into multiple outlets. (b) Possible microfluidic channel configurations for density-based separation: Uniform channel height (left), gradual channel expansion (middle), and hydraulic jump cavity with sudden channel expansion (right). The height difference between particles with different densities can be amplified by the sudden channel expansion compared to the other two cases due to the relatively large tangential angle, θ of F_D. (|θ₁|≪ |θ₂|) (see Fig. S1 in the supplementary material²²).In comparison with the other two cases (Fig. 1(b) uniform channel height and gradual channel expansion), the height difference between the particles with different densities can be amplified by the sudden channel expansion in the hydraulic jump cavity due to relatively large tangential angle (see supplementary material²²). Therefore, the particles can be separated through the multiple outlets, depending on their height and distance.In order to analyze the separation behavior of particles in the chamber according to differences in their densities, H and D are systematically investigated. The numerical simulations are performed using a commercial CFD software (CFX 14.0; ANSYS 14.0; ANSYS, Inc.). Particles with the same density may have different trajectories in the separation chamber depending on their inlet positions (Fig. 2(a)). Prior to this investigation, the maximum height (H_max) and distance (D_max) for each particle are compared by examining H and D of 100 identical particles at different inlet positions since the inlet position of particles could be controlled.²⁰ Fig. 2(b) shows H_max and D_max of particles with respect to density at a fixed Reynolds number (Re = 0.1). Note that Reynolds number is defined as Re = ρ_fv_fD_h/μ, where ρ_f, v_f, D_h, μ are density of fluid, velocity of the fluid, hydraulic diameter of a channel, and dynamic viscosity of the fluid, respectively. The hydraulic diameter in the Reynolds number is determined with the inlet channel. Particle densities in the range of 1.1 to 2.0 g/cm³ are chosen with the increase of 0.1 g/cm³. These values are quite reasonable in that the densities of many microorganisms such as microalgae are typically within this range and their densities can be varied by 0.2 g/m³ depending on their cellular context.²³ The lighter particles travel with a higher H_max, and longer D_max. With the separation chamber, the height difference between particles with densities of 1.1 and 1.2 g/cm³ can be amplified by about 10 times as compared to that in a channel without the chamber, judging from the position where the 1.1 g/cm³ particle reaches its H_max.Open in a separate windowFIG. 2.Microfluidic particle separation with respect to Reynolds number (Re). (a) Trajectories in the separation chamber of a hundred particles with the same density starting from inlet positions chosen arbitrarily in order to investigate the effect of the inlet positions on the maxima of the height (H_max) and distance (D_max) prior to further simulation. (b) Representative trajectories of particles having different densities from 1.1 to 2.0 g/cm³. (c) The maximum height (H_max) of each particle with respect to Re. (d) Representative maximum distance (D_max) of each particle at Re = 0.1. (Left) Streamline of fluid and representative trajectories of particles with densities of 1.1 and 2.0 g/cm³ in the separation chamber at Re = 0.1 (right).In Fig. 2(c), the values for H_max of particles with respect to Reynolds number (Re) are presented. Since in our study, the maximum height (H_max) and distance (D_max) for each particle were compared by examining H and D of 100 identical particles that are randomly distributed in the channel (throughout all figures), there is little variation in H_max and D_max between each simulation. However, the standard deviation between each simulation is quite small and can be negligible. The H_max values particles at Re = 0.5 with densities of 1.1 g/cm³ and 1.2 g/cm³ are 2.21 × 10³μm and 2.17 × 10³μm, respectively. The difference between H_max of different particles, ΔH_max, increases with decreasing Re. For example, ΔH_max between particles with densities of 1.1 and 2.0 g/cm³ becomes 0.26 × 10³μm at Re = 1.0, but increases to 1.38 × 10³μm as Re decreases to 0.1. As Re increases (velocity of fluid increases), the relative velocity in the y direction between the fluid and the particle increases resulting in increasing of F_D in the y direction since the velocity of particle in the y direction is very small at the entrance of the separation chamber. Thus, contribution of F_D becomes comparable to the net force in the y direction. As a result, most of the particles even in the case of heavier ones travel quite similarly with the streamline, and ΔH_max subsequently decreases. On the other hand, as Re decreases, the contribution of F_G becomes dominant due to the decrease of F_D in the y direction. Consequently, the particles start to cross downwards streamlines as the density of the particles increases and H_max gradually decreases. In addition, irrespective of their densities, ΔH_max of the particles increases with decreasing Re.Fig. 2(d) shows D_max with respect to the density of the particles (left). Different densities of particles show different trajectories due to the relative contribution of F_D to the net force in the y direction depending on the particle density (right). At Re = 0.1, D_max of particles with densities of 1.1 cm³ and 1.2 g/cm³ are 2.91 × 10⁴μm and 1.43 × 10⁴μm, respectively. As the density of a particle increases, its D_max dramatically decreases. The difference in D_max between particles with densities of 1.1 and 1.2 g/cm³ is 1.48 × 10⁴μm, and 0.0037 × 10⁴μm for particles with densities of 1.9 and 2.0 g/cm³. The effect of F_D is stronger compared to that of F_G on lighter particles. Thus, lighter particles travel quite similarly with the streamline and finally have a large D_max. On the other hand, heavier particles where effect of F_G is stronger compared to that of F_D cross downwards streamlines and finally have a small D_max.Next, in order to investigate the separation behavior of particles with respect to the geometry of the microfluidic device, the effect of the ratio of the height of the separation chamber (h_c) to the inlet (h_i) on H_max is investigated as shown in Fig. ​Fig.3.3. Interestingly, H_max of particles with density of 1.1 g/cm³ increases from 1.93 × 10³μm to 6.48 × 10³μm while that of particles with density of 1.9 g/cm³ slightly changes from 0.70 × 10³μm to 0.73 × 10³μm as h_c/h_i increases from 5 to 20.Open in a separate windowFIG. 3.Microfluidic particle separation with respect to the ratio of the height of the inlet (h_i) to the separation chamber (h_c).This result can be attributed to two effects: (1) the change in the streamline and (2) the relative contribution of drag force to the net force depending on the density. With increasing h_c/h_i, dramatic increase in H_max for lighter particles is because the streamline for the lighter ones experiences more vertical displacement in the separation chamber and the contribution of F_D to the net force acting on the lighter one is more significant (see Fig. S2 in the supplementary material²²).Based on this approach, we propose a microfluidic device for the selective separation of the lightest particle. Fig. 4(a) shows one unit (with three outlets) of the proposed microfluidic device that can be connected in series. The ratio of channel heights (h_c/h_i) is set to 20, and the particle densities are in the range of 1.1 ∼ 1.5 g/m³. Fig. 4(b) shows the representative separation behavior of the particles. A portion of the lightest particles (1.1 g/cm³) is selectively separated into the upper and middle outlets, while remaining light particles together with four other heavier particles with densities in the range of 1.2 to 1.5 g/cm³ leave through the lowest outlet. With a single operation of this unit, 40% of the lightest particles are recovered. In addition, the yield increases with increasing number of cycles (Fig. 4(c)).Open in a separate windowFIG. 4.(a) One unit of the proposed microfluidic device for the selective separation of the lightest particle based on the simulation results. Particles are separated into two outlets based on differences in both the height and distance travelled stemming from differences in density. (b) Representative separation behavior of particles observed in the device. (c) The yield of the lightest particle (1.1 g/cm³) with the proposed microfluidic device according to the number of cycles (i.e., this unit is assumed to be connected in series).In summary, we have demonstrated a label-free microfluidic system for the separation of particles according to subtle differences in their densities without external forces. Our microfluidic design consists simply of an inlet, a separation chamber, and multiple outlets. When entering the separation chamber, the particles experience an additional drag force in the y direction, amplifying the difference in both the height and the distance that the particles with different densities can travel within the chamber. At a fixed Reynolds number, with increasing particle density, H_max decreases monotonously, and D_max decreases dramatically. On the other hand, as Reynolds number increases, the difference between the heights of particles with different densities is attenuated. In addition, the simulation reveals that increasing the ratio of the channel heights increases the difference between the heights of particles only when their densities are close to that of the surrounding water. Based on this approach, a microfluidic device for the separation of the lightest particles has been proposed. We expect that our density-based separation design can be beneficial to the selective separation of specific microorganisms such as lipid-rich microalgae for energy harvesting application.     

Synoptic curriculum texts: representation of contemporary curriculum scholarship

Building upon past studies of curriculum textbooks in the US, this paper presents an analytical framework that classifies curriculum texts according to their representation of reconceptualized curriculum scholarship. It uses four indicators of the representation of curriculum scholarship in the analyses of eight curriculum texts published in the USA in the 1990s: the author’s stated intentions; the organization of the table of contents; the treatment of the forms of reconceptualism; and the ratio of curriculum and non‐curriculum journal citations. The analysis finds that reconceptualized curriculum scholarship has been at least mentioned, if not extensively treated, in most texts since the mid‐1980s, and that this approach is most comprehensively treated in books published in the 1990s.     

Assessment and Learning of Qualitative Physics in Newton's Playground

ABSTRACT

Digital games are very popular in modern culture. The authors are examining ways to leverage these engaging environments to assess and support student competencies. The authors examine gameplay and learning using a physics game they developed called Newton's Playground. The sample consisted of 167 eighth- and ninth-grade students who played Newton's Playground for about 4 hr over the course of 1.5 weeks. Findings include significant pretest–posttest physics gains, and significant relations between in-game indicators and learning.     

Social alienation by communication denial

The major claim of this essay is that an important relationship between hermeneutics and rhetoric has been overlooked by communication scholars. By describing the nature of this relationship the authors clarify rhetoric's epistemic function and, in turn, provide important theoretical directives for rhetorical criticism.     

MEMORY,SELF‐EFFICACY,AND ADAPTABILITY IN KOREAN AMERICAN OLDER ADULTS: A COLLECTIVE STUDY OF FOUR CASES

Currently influential educational theories emphasize a causal and linear relationship among memory self‐efficacy, psychological well‐being, and memory performance. That model, however, does not consider an important mediating variable: the adaptability of the aging self. This collective study of four cases (four Korean American older adults) was conducted to provide a more comprehensive understanding of the interrelationships among these variables, with the expectation that useful insights might be gained through the in‐depth study of specific cases. Each case showed a unique pattern of the interplay of these cognitive and affective variables. The findings suggest that perceptions of memory functioning and psychological well‐being are mitigated by the adaptability of the aging self, rather than being directly related to the experiences of age‐related losses or to actual memory perfor‐     

MOTIVATIONS FOR LEARNING AMONG OLDER ADULTS IN A LEARNING IN RETIREMENT INSTITUTE

As the population of older adults increases, the field of adult education needs to respond accordingly. The study reported in this paper examined motivations for learning among older adults actively engaged in formal lifelong learning. One hundred eighty-nine members of a Learning in Retirement institute were surveyed using Boshier's Education Participation Scale. Cognitive interest appeared to be the strongest motivator for learning. Social contact was the second most influential motivator. The findings of this study are consistent with, and add to, our knowledge derived from previous studies pertaining to the participation of older adults in formal learning. Recommendations for future research on older adult learning in different settings are presented.     

Reviewing and Critiquing Computer Learning and Usage Among Older Adults

By searching the keywords of “older adult” and “computer” in ERIC, Academic Search Premier, and PsycINFO, this study reviewed 70 studies published after 1990 that address older adults' computer learning and usage. This study revealed 5 prominent themes among reviewed literature: (a) motivations and barriers of older adults' usage of computers, (b) age-related differences in computer learning and usage, (c) instructional tips and design, (d) changes in attitudes and benefits, and (e) Internet usage. The limitations of the studies are reviewed and suggestions for future research are discussed.     

Labour market motivation and undergraduates' choice of degree subject

Labour market outcomes of undergraduates' choice of subject are important for public policy and for students. Policy interest is indicated by the prominence of ‘employability’ in public discourse and in proposals to concentrate government funding in England in supporting STEM subjects (science, technology, engineering and mathematics). As students in England face the prospect of bearing the full financial burden of undergraduate tuition, the large differences between wage premia for different subjects may become of increasing interest. We find that, even after taking account of differences in motivation towards the choice of undergraduate subject, males and members of certain non‐White ethnic groups are more likely to choose ‘high wage‐premium’ subjects. We also find some significant differences between the motivations of different minority ethnic groups. However, students from lower income households are less likely to choose high wage premium subjects, which is a concern for this aspect of policy towards participation in higher education and social mobility.     

An investigation of the effects of a graphic organizer in an online serious game on learning outcomes and attitudinal perceptions

A serious game, which is designed for learning purposes rather than recreational purposes, has been applied for digital game-based Learning. This study investigated the effects of graphic organizers in a serious game, “The Transistor”, on learning outcomes and attitudinal perceptions. A total of 99 participants were randomly assigned to three groups: non-organizer group, advance organizer group, and post organizer group. The results revealed that participants using a post organizer, which was presented after the game for review, outperformed those using a non-organizer and an advance organizer on both learning outcomes (recall test scores and recycling intention) and attitudinal perceptions (perceived enjoyment, perceived usefulness, and satisfaction). These findings implied that a post-graphic organizer could enhance the instructional value of a serious game. More detailed implications, limitations, and suggestions for future research are discussed.     

Expanding “within context” to “across contexts” learning: a case study of informal and formal activities

Learning happens not only in schools, but also in every context that affords new experiences and opportunities for metacognition. We aim to maximize the different activity-milieux in which learners are engaged in developing their life-long learning dispositions to learn within and across contexts. This paper is a follow up of an earlier published paper in which a framework on interpreting learning as how the self interacts with phenomena and reifications was proposed. Grounded in this learning framework, our research seeks to expand the “within context” learning to “across contexts” learning with evidence from a case study examining a 10-year-old boy's learning process. Through the case study, we will describe the interplay between bowling and schooling and how strategies learned from one context is situated into another drawing upon Ito et al.'s (2010 Ito, M., Baumer, S., Bittanti, M, boyd, d., Cody, R., Herr-Stephenson, B., Horst, … Pascoe, C.J., Perkel, D., Robinson, L., Sims, C., Tripp, L. (2010). Hanging out, messing around, and geeking out: Kids living and learning with new media. Cambridge, MA: MIT Press [Google Scholar]) developmental trajectory of “hanging out-messing around-geeking out” perspectives. The findings show that for learning to be meaningful and authentic for the twenty-first century, it is essential for the learner(s) to be in constant dialectical interactions and relationship with oneself, others, and with the social-cultural artifacts that afford the learning. We conclude with practical implications derived from the study.