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Methods in Cell Biology

Junsang Doh, Daniel A Fletcher, Matthieu Piel
No abstract text is available yet for this article.
2018: Methods in Cell Biology
Sinan Muldur, Anika L Marand, Felix Ellett, Daniel Irimia
Neutrophils play an essential role in the protection against infection, as they are the most numerous circulating white blood cell population and the first responders to injury. Their numbers in blood are frequently measured in the clinic and used as an indicator of ongoing infections. During inflammation and sepsis, the ability of neutrophils to migrate is disrupted, which may increase the risk of infection, even when the neutrophil count is normal. However, measurements of neutrophil migration in patients are rarely performed because of the challenges of performing the migration assays in a clinical setting...
2018: Methods in Cell Biology
Jörg Renkawitz, Anne Reversat, Alex Leithner, Jack Merrin, Michael Sixt
Cells migrating in multicellular organisms steadily traverse complex three-dimensional (3D) environments. To decipher the underlying cell biology, current experimental setups either use simplified 2D, tissue-mimetic 3D (e.g., collagen matrices) or in vivo environments. While only in vivo experiments are truly physiological, they do not allow for precise manipulation of environmental parameters. 2D in vitro experiments do allow mechanical and chemical manipulations, but increasing evidence demonstrates substantial differences of migratory mechanisms in 2D and 3D...
2018: Methods in Cell Biology
Lucile Alexandre, Iago Pereiro, Amel Bendali, Sanae Tabnaoui, Jana Srbova, Zuzana Bilkova, Shane Deegan, Lokesh Joshi, Jean-Louis Viovy, Laurent Malaquin, Bruno Dupuy, Stéphanie Descroix
Bacterial contamination and subsequent infections are a major threat to human health. An early detection in the food chain, clinics or the environment, is key to limit this threat. We present a new concept to develop low-cost hand-held devices for the ultra-sensitive and specific detection of bacteria in a one-step process of 2-8h, directly from complex raw samples. This approach is based on a novel microfluidic magnetic fluidized bed. It reaches a 4CFU (colony forming unit) sensitivity with high quantification accuracy in a large dynamic range of 100-107 CFU/mL...
2018: Methods in Cell Biology
Julia Bos, Robert H Austin
The systematic emergence of drug resistance remains a major problem in the treatment of infectious diseases (antibiotics) and cancer (chemotherapy), with possible common fundamental origins linking bacterial antibiotic resistance and emergence of chemotherapy resistance. The common link may be evolution in a complex fitness landscape with connected small population niches. We report a detailed method for observing bacterial adaptive behavior in heterogeneous microfluidic environment designed to mimic the environmental heterogeneity found in natural microbial niches...
2018: Methods in Cell Biology
Javier Muñoz-Garcia, Julien Babic, Damien Coudreuse
Microfluidic technologies have become a standard tool in cell biological studies, offering unprecedented control of the chemical and physical environment of cells grown in microdevices, the possibility of multiplexing assays, as well as the capacity to monitor the behavior of single cells in real time while dynamically manipulating their growth medium. However, the properties of the materials employed for the fabrication of microchips that are compatible with live-cell imaging has limited the use of these techniques for a broad range of experiments...
2018: Methods in Cell Biology
Youlian Goulev, Audrey Matifas, Gilles Charvin
The generation of complex temporal stress patterns may be instrumental to investigate the adaptive properties of individual cells submitted to environmental stress on physiological timescale. However, it is difficult to accurately control stress concentration over time in bulk experiments. Here, we describe a microfluidics-based protocol to induce tightly controllable H2 O2 stress in budding yeast while constantly monitoring cell growth with single cell resolution over multi-generation timescale. Moreover, we describe a simple methodology to produce ramping H2 O2 stress to investigate the homeostatic properties of the H2 O2 scavenging system...
2018: Methods in Cell Biology
L J Holt, O Hallatschek, M Delarue
Cells need to act upon the elastic extracellular matrix and against steric constraints when proliferating in a confined environment, leading to the build-up, at the population level, of a compressive, growth-induced, mechanical stress. Compressive mechanical stresses are ubiquitous to any cell population growing in a spatially-constrained environment, such as microbes or most solid tumors. They remain understudied, in particular in microbial populations, due to the lack of tools available to researchers. Here, we present various mechano-chemostats: microfluidic devices developed to study microbes under pressure...
2018: Methods in Cell Biology
Chiara Toniolo, Matthieu Delincé, John D McKinney
Single-cell imaging of host-microbe interactions over time is impeded by cellular motility because the cells under scrutiny tend to migrate out of the imaging field. To overcome this technical challenge, we developed a microfluidic platform for imaging hundreds of individual motile phagocytic cells and bacteria within microfluidic traps that restrict their movement. The interaction of trapped host cells and bacteria is monitored by long-term time-lapse microscopy, allowing direct visualization of all stages of infection at the single-cell level...
2018: Methods in Cell Biology
Marta Urbanska, Philipp Rosendahl, Martin Kräter, Jochen Guck
Mechanical properties of cells can serve as a label-free marker of cell state and function and their alterations have been implicated in processes such as cancer metastasis, leukocyte activation, or stem cell differentiation. Over recent years, new techniques for single-cell mechanical characterization at high throughput have been developed to accelerate discovery in the field of mechanical phenotyping. One such technique is real-time deformability cytometry (RT-DC), a robust technology based on microfluidics that performs continuous mechanical characterization of cells in a contactless manner at rates of up to 1000 cells per second...
2018: Methods in Cell Biology
Ramanathan Vaidyanathan, Trifanny Yeo, Chwee Teck Lim
The complexity and dynamic evolution of cancer often result in tumor subpopulations containing distinctly heterogeneous cells. During metastasis, these also give rise to heterogeneous circulating tumor cells (CTCs) which are considered to be a hematogenous dissemination from the primary tumor. CTCs represent a viable less-invasive sampling opportunity, also known as liquid biopsy. However, current technological platforms that analyze entire CTC population are not effective due to cell-to-cell variability within the same population and this can manifest differences in genomic expression, cell cycle stages and eventually cellular responses to drug treatments...
2018: Methods in Cell Biology
Andrew G Clark, Anthony Simon, Koceila Aizel, Jérôme Bibette, Nicolas Bremond, Danijela Matic Vignjevic
Chemotaxis is an important biological process involved in the development of multicellular organisms, immune response and cancer metastasis. In order to better understand how cells follow chemical cues in their native environments, we recently developed a microfluidics-based chemotaxis device that allows for observation of cells or cell aggregates in 3D networks in response to tunable chemical gradients (Aizel et al., 2017). Here, we describe the methods required for fabrication of this device as well as its use for live imaging experiments and subsequent analysis of imaging data...
2018: Methods in Cell Biology
Emilie Le Maout, Simon Lo Vecchio, Alka Bhat, Daniel Riveline
Cell motility has been mainly characterized in vitro through the motion of cells on 2D flat Petri dishes, and in Boyden chambers with the passage of cells through sub-cellular sized cavities. These experimental conditions have contributed to understand important features, but these artificial designs can prevent elucidation of mechanisms involved in guiding cell migration in vivo. In this context, microfabrication and microfluidics have provided unprecedented tools to design new assays with local controls in two and three dimensions...
2018: Methods in Cell Biology
Junsang Doh, Daniel A Fletcher, Matthieu Piel
No abstract text is available yet for this article.
2018: Methods in Cell Biology
Jeonghwan Lee, Kipyo Kim, Sejoong Kim
Kidney tubule-on-a-chip has been developed since 2001. Drug toxicity and pharmacokinetic models using kidney tubule-on-a-chip devices are already available. Experimental models for other causes of acute kidney injury such as ischemia or infection will be available in the near future. Recently, glomerulus-on-a-chip was introduced using induced pluripotent stem cells. It is a far advanced model using the glomerulus structure. Its applications are still challenging but promising since patients with chronic kidney disease are steadily increasing with the increased prevalence of diabetes, hypertension, and old age...
2018: Methods in Cell Biology
Giovanni Stefano Ugolini, Roberta Visone, Daniela Cruz-Moreira, Andrea Mainardi, Marco Rasponi
With the increasing attention on cardiovascular disorders and the current inability of pre-clinical models to accurately predict human physiology, the need for advanced and reliable heart in vitro models is paramount. Microfabrication technologies provide potential solutions in the organs-on-chip systems: microengineered devices where cell cultures can be hosted and cultured to develop three-dimensional models or microtissues with high similarity to human physiology. We here described the fabrication and operation procedures for a beating heart-on-a-chip...
2018: Methods in Cell Biology
Fan Lee, Ciprian Iliescu, Fang Yu, Hanry Yu
3D spheroid or organoid culture is becoming mainstream method for studying physiologically relevant cell behaviors, and used in applications such as cell-based diagnostic, therapy, disease modeling and drug screening. Organoids/spheroids function best when maintaining size of <200μm diameter and in perfusion culture. To achieve this, we describe in this chapter various methods of constraining spheroid size during the formation and culture processes such that the spheroids can maintain high-level cell functions for characterization and applications...
2018: Methods in Cell Biology
Wang Xi, Surabhi Sonam, Chwee Teck Lim, Benoit Ladoux
Epithelial cells demonstrate different collective migratory modes when encountering two (2D) and three dimensional (3D) microenvironment. While planar micropatterns and constraint have been shown to strongly impact collective cell migration (CCM), how out-of-plane curvature and 3D confinement will affect epithelial organization and dynamics remains largely unknown. This is likely due to lack of proper 3D microscaffolds for studying CCM. In this chapter, we briefly review the latest achievement in microengineering approaches to control 3D microenvironment of epithelial development...
2018: Methods in Cell Biology
Jaehyun Lee, Hyung Kyu Huh, Sung Ho Park, Sang Joon Lee, Junsang Doh
In response to inflammatory signals, leukocytes circulating in blood vessels undergo dynamic interaction with endothelium lining blood vessel walls, known as leukocyte adhesion cascade, to leave the blood vessels, and infiltrate the inflamed tissue. During leukocyte extravasation, leukocytes recognize and respond to various biophysical and biochemical cues present in the complex microenvironments of inflamed blood vessels to find optimal pathways. Although advances in intravital imaging of live animals have enabled us to observe leukocyte dynamics during extravasation, in vitro model systems mimicking complex in vivo microenvironments are still needed for mechanistic studies...
2018: Methods in Cell Biology
Anna A Kim, Adam L Nekimken, Sylvia Fechner, Lucy E O'Brien, Beth L Pruitt
Mechanical stimuli play a critical role in organ development, tissue homeostasis, and disease. Understanding how mechanical signals are processed in multicellular model systems is critical for connecting cellular processes to tissue- and organism-level responses. However, progress in the field that studies these phenomena, mechanobiology, has been limited by lack of appropriate experimental techniques for applying repeatable mechanical stimuli to intact organs and model organisms. Microfluidic platforms, a subgroup of microsystems that use liquid flow for manipulation of objects, are a promising tool for studying mechanobiology of small model organisms due to their size scale and ease of customization...
2018: Methods in Cell Biology
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