journal
MENU ▼
Read by QxMD icon Read
search

Current Protocols in Cell Biology

journal
https://www.readbyqxmd.com/read/28862343/cellular-detection-of-g-quadruplexes-by-optical-imaging-methods
#1
Souheila Amor, Sunny Y Yang, Judy M Y Wong, David Monchaud
G-quadruplexes (G4s) are higher-order nucleic acid structures that fold from guanine (G)-rich DNA and RNA strands. This field of research gains traction as a major chemical biology area since it aims at uncovering many key cellular mechanisms in which quadruplexes are involved. The wealth of knowledge acquired over the past three decades strongly supports pivotal roles of G4 in the regulation of gene expression at both transcriptional (DNA quadruplexes) and translational levels (RNA quadruplexes). Recent biochemical discoveries uncovered myriad of additional G4 actions: from chromosomal stability to the firing of replication origins, from telomere homeostasis to functional dysregulations underlying genetic diseases (including cancers and neurodegeneration)...
September 1, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28862342/lectin-array-blotting
#2
Raquel Pazos, Juan Echevarria, Alvaro Hernandez, Niels-Christian Reichardt
Aberrant protein glycosylation is a hallmark of cancer, infectious diseases, and autoimmune or neurodegenerative disorders. Unlocking the potential of glycans as disease markers will require rapid and unbiased glycoproteomics methods for glycan biomarker discovery. The present method is a facile and rapid protocol for qualitative analysis of protein glycosylation in complex biological mixtures. While traditional lectin arrays only provide an average signal for the glycans in the mixture, which is usually dominated by the most abundant proteins, our method provides individual lectin binding profiles for all proteins separated in the gel electrophoresis step...
September 1, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28862341/centrifugation-free-magnetic-isolation-of-functional-mitochondria-using-paramagnetic-iron-oxide-nanoparticles
#3
Bhabatosh Banik, Shanta Dhar
Subcellular fractionation techniques are essential for cell biology and drug development studies. The emergence of organelle-targeted nanoparticle (NP) platforms necessitates the isolation of target organelles to study drug delivery and activity. Mitochondria-targeted NPs have attracted the attention of researchers around the globe, since mitochondrial dysfunctions can cause a wide range of diseases. Conventional mitochondria isolation methods involve high-speed centrifugation. The problem with high-speed centrifugation-based isolation of NP-loaded mitochondria is that NPs can pellet even if they are not bound to mitochondria...
September 1, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28862340/in-vitro-reconstitution-of-the-endoplasmic-reticulum
#4
Csilla-Maria Ferencz, Gernot Guigas, Andreas Veres, Brigitte Neumann, Olaf Stemmann, Matthias Weiss
Reconstitution of cellular organelles in vitro offers the possibility to perform quantitative and qualitative experiments in a controlled environment that cannot be done with the same accuracy in living cells. Following a previous report, the subsequent list of protocols describes how to reconstitute and quantify a tubular ER network in vitro based on purified microsomes from culture cells and cytosol from Xenopus laevis egg extracts. Biological material preparation and reconstitution assays require mostly basic laboratory instrumentation and chemicals, and can be executed without any specific training, making them appealing to a wide range of laboratories...
September 1, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28862339/determination-of-membrane-protein-distribution-on-the-nuclear-envelope-by-single-point-single-molecule-frap
#5
Krishna C Mudumbi, Weidong Yang
Nuclear envelope transmembrane proteins (NETs) are synthesized on the endoplasmic reticulum and then transported from the outer nuclear membrane (ONM) to the inner nuclear membrane (INM) in eukaryotic cells. The abnormal distribution of NETs has been associated with many human diseases. However, quantitative determination of the spatial distribution and translocation dynamics of NETs on the ONM and INM is still very limited in currently existing approaches. Here we demonstrate a single-point single-molecule fluorescence recovery after photobleaching (FRAP) microscopy technique that enables quick determination of distribution and translocation rates for NETs in vivo...
September 1, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28862338/automated-tracking-of-cell-migration-with-rapid-data-analysis
#6
Brian J DuChez
Cell migration is essential for many biological processes including development, wound healing, and metastasis. However, studying cell migration often requires the time-consuming and labor-intensive task of manually tracking cells. To accelerate the task of obtaining coordinate positions of migrating cells, we have developed a graphical user interface (GUI) capable of automating the tracking of fluorescently labeled nuclei. This GUI provides an intuitive user interface that makes automated tracking accessible to researchers with no image-processing experience or familiarity with particle-tracking approaches...
September 1, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28627757/super-resolution-microscopy-and-single-protein-tracking-in-live-bacteria-using-a-genetically-encoded-photostable-fluoromodule
#7
Saumya Saurabh, Adam M Perez, Colin J Comerci, Lucy Shapiro, W E Moerner
Visualization of dynamic protein structures in live cells is crucial for understanding the mechanisms governing biological processes. Fluorescence microscopy is a sensitive tool for this purpose. In order to image proteins in live bacteria using fluorescence microscopy, one typically genetically fuses the protein of interest to a photostable fluorescent tag. Several labeling schemes are available to accomplish this. Particularly, hybrid tags that combine a fluorescent or fluorogenic dye with a genetically encoded protein (such as enzymatic labels) have been used successfully in multiple cell types...
June 19, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28627756/microcontact-peeling-a-cell-micropatterning-technique-for-circumventing-direct-adsorption-of-proteins-to-hydrophobic-pdms
#8
Sho Yokoyama, Tsubasa S Matsui, Shinji Deguchi
Microcontact printing (μCPr) is one of the most popular techniques used for cell micropatterning. In conventional μCPr, a polydimethylsiloxane (PDMS) stamp with microfeatures is used to adsorb extracellular matrix (ECM) proteins onto the featured surface and transfer them onto particular areas of a cell culture substrate. However, some types of functional proteins other than ECM have been reported to denature upon direct adsorption to hydrophobic PDMS. Here we describe a detailed protocol of an alternative technique--microcontact peeling (μCPe)--that allows for cell micropatterning while circumventing the step of adsorbing proteins to bare PDMS...
June 19, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28627755/labeling-dna-replication-foci-to-visualize-chromosome-territories-in-vivo
#9
Apolinar Maya-Mendoza, Dean A Jackson
While a detailed understanding of chromatin dynamics is needed to explain how higher-order chromatin organization influences nuclear function, the molecular principles that regulate chromatin mobility in mammalian nuclei remain largely unknown. Here we describe experimental tools to follow chromatin dynamics by labeling DNA during S phase. Using these methods, we have found that foci labeled during early and mid/late S phase have significantly different dynamic behavior. Spatially constrained heterochromatic foci restrict long-range transformations of the chromosome territory (CT) structure while providing a structural framework on which highly mobile euchromatic foci undergo positional oscillations that drive local changes in the chromosome shape...
June 19, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28627754/labeling-and-magnetic-resonance-imaging-of-exosomes-isolated-from-adipose-stem-cells
#10
Alice Busato, Roberta Bonafede, Pietro Bontempi, Ilaria Scambi, Lorenzo Schiaffino, Donatella Benati, Manuela Malatesta, Andrea Sbarbati, Pasquina Marzola, Raffaella Mariotti
Adipose stem cells (ASC) represent a promising therapeutic approach for neurodegenerative diseases. Most biological effects of ASC are probably mediated by extracellular vesicles, such as exosomes, which influence the surrounding cells. Current development of exosome therapies requires efficient and noninvasive methods to localize, monitor, and track the exosomes. Among imaging methods used for this purpose, magnetic resonance imaging (MRI) has advantages: high spatial resolution, rapid in vivo acquisition, and radiation-free operation...
June 19, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28627753/traction-force-microscopy-in-3-dimensional-extracellular-matrix-networks
#11
M Cóndor, J Steinwachs, C Mark, J M García-Aznar, B Fabry
Cell migration through a three-dimensional (3-D) matrix depends strongly on the ability of cells to generate traction forces. To overcome the steric hindrance of the matrix, cells need to generate sufficiently high traction forces but also need to distribute these forces spatially in a migration-promoting way. This unit describes a protocol to measure spatial maps of cell traction forces in 3-D biopolymer networks such as collagen, fibrin, or Matrigel. Traction forces are computed from the relationship between measured force-induced matrix deformations surrounding the cell and the known mechanical properties of the matrix...
June 19, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28627752/patterning-on-topography-for-generation-of-cell-culture-substrates-with-independent-nanoscale-control-of-chemical-and-topographical-extracellular-matrix-cues
#12
Emily N Sevcik, John M Szymanski, Quentin Jallerat, Adam W Feinberg
The cell microenvironment plays an important role in many biological processes, including development and disease progression. Key to this is the extracellular matrix (ECM), a complex biopolymer network serving as the primary insoluble signaling network for physical, chemical, and mechanical cues. In vitro, the ability to engineer the ECM at the micro- and nanoscales is a critical tool to systematically interrogate the influence of ECM properties on cellular responses. Specifically, both topographical and chemical surface patterning has been shown to direct cell alignment and tissue architecture on biomaterial surfaces, however, it has proven challenging to independently control these surface properties...
June 19, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28256723/immunoblotting-and-immunodetection
#13
Duojiao Ni, Peng Xu, Sean Gallagher
Immunoblotting (western blotting) is used to identify specific antigens recognized by polyclonal or monoclonal antibodies. This unit provides protocols for all steps, starting with solubilization of the protein samples, usually by means of SDS and reducing agents. Following solubilization, the material is separated by SDS-PAGE and the antigens are electrophoretically transferred to a membrane, a process that can be monitored by reversible staining with Ponceau S. The transferred proteins are bound to the surface of the membrane, providing access to immunodetection reagents...
March 3, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28256722/discovering-protein-protein-interactions-using-nucleic-acid-programmable-protein-arrays
#14
Yanyang Tang, Ji Qiu, Matthias Machner, Joshua LaBaer
We have developed a protocol enabling the study of protein-protein interactions (PPIs) at the proteome level using in vitro-synthesized proteins. Assay preparation requires molecular cloning of the query gene into a vector that supports in vitro transcription/translation (IVTT) and appends a HaloTag to the query protein of interest. In parallel, protein microarrays are prepared by printing plasmids encoding glutathione S-transferase (GST)-tagged target proteins onto a carrier matrix/glass slide coated with antibody directed against GST...
March 3, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28256721/a-functional-microrna-screening-method-for-organ-morphogenesis
#15
Ivan T Rebustini
The increasing repertoire of microRNAs expressed during organ development and their role in regulating organ morphogenesis provide a compelling need to develop methods to assess microRNA function using various in vitro and in vivo experimental models. Methods to assess microRNA function during organ morphogenesis include transfection of microRNA inhibitors (antagomirs) and activators (mimics) into mouse embryonic explanted organs using liposomes, which can potentially result in low efficiency of transfection and off-target effects...
March 3, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28256720/a-unique-high-throughput-assay-to-identify-novel-small-molecule-inhibitors-of-chemotaxis-and-migration
#16
Xin-Hua Liao, Alan R Kimmel
Chemotaxis and cell migration play pivotal roles in normal physiological processes such as embryogenesis, inflammation, and wound healing, as well as in pathological processes including chronic inflammatory disease and cancer metastasis. Novel chemotaxis/migration inhibitors are desirable for developing effective therapeutics and probing molecular mechanisms. We describe a fluorescence-based phenotypic assay in a 1536-well plate format for high-throughput screening of novel inhibitors of chemotaxis/migration within complex libraries of thousands of compounds...
March 3, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28256719/rna-whole-mount-in-situ-hybridization-proximity-ligation-assay-rish-pla-an-assay-for-detecting-rna-protein-complexes-in-intact-cells
#17
Ioannis M Roussis, Fiona A Myers, Garry P Scarlett
Techniques for studying RNA-protein interactions have lagged behind those for DNA-protein interactions as a consequence of the complexities associated with working with RNA. This unit describes a method for the adaptation of the In Situ Hybridization-Proximity Ligation Assay (ISH-PLA) to the study of RNA regulation (rISH-PLA). The rISH-PLA assay allows the identification of a given RNA-protein complex at subcellular and single-cell resolution, thus avoiding the lack of spatial resolution and sensitivity associated with assaying heterogeneous cell populations from which conventional RNA-protein interaction detection techniques suffer...
March 3, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/27906451/proximity-dependent-biotinylation-for-identification-of-interacting-proteins
#18
Valerie Le Sage, Alessandro Cinti, Andrew J Mouland
Complex interaction networks orchestrate key cellular processes including but not limited to transcription, translation, metabolism, and cell signaling. Delineating these interactions will aid in deciphering the regulation and function of these pathways and potential for manipulation. Proximity-dependent biotin identification (BioID) is quickly gaining popularity as a powerful tool for identifying novel protein-protein and proximity-based interactions in live cells. This technique relies on a promiscuous biotin ligase, which is fused to a protein of interest and, upon expression in the desired cell, will biotinylate proximal endogenous proteins...
December 1, 2016: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/27906450/lovtrap-a-versatile-method-to-control-protein-function-with-light
#19
Hui Wang, Klaus M Hahn
We describe a detailed procedure for the use of LOVTRAP, an approach to reversibly sequester and release proteins from cellular membranes using light. In the application described here, proteins that act at the plasma membrane are held at mitochondria in the dark, and reversibly released by irradiation. The technique relies on binding of an engineered Zdk domain to a LOV2 domain, with affinity <30 nM in the dark and >500 nM upon irradiation between 400 and 500 nm. LOVTRAP can be applied to diverse proteins, as it requires attaching only one member of the Zdk/LOV2 pair to the target protein, and the other to the membrane where the target protein is to be sequestered...
December 1, 2016: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/27580706/isolation-of-lipid-droplets-from-cells-by-density-gradient-centrifugation
#20
Dawn L Brasaemle, Nathan E Wolins
Lipid droplets are organelles found in most mammalian cells, as well as in various plant tissues and yeast. They are composed of a core of neutral lipids surrounded by a membrane monolayer of phospholipids and cholesterol in which specific proteins are embedded. This unit provides protocols for isolating lipid droplets from mammalian cells by discontinuous density gradient centrifugation. © 2016 by John Wiley & Sons, Inc.
September 1, 2016: Current Protocols in Cell Biology
journal
journal
41482
1
2
Fetch more papers »
Fetching more papers... Fetching...
Read by QxMD. Sign in or create an account to discover new knowledge that matter to you.
Remove bar
Read by QxMD icon Read
×

Search Tips

Use Boolean operators: AND/OR

diabetic AND foot
diabetes OR diabetic

Exclude a word using the 'minus' sign

Virchow -triad

Use Parentheses

water AND (cup OR glass)

Add an asterisk (*) at end of a word to include word stems

Neuro* will search for Neurology, Neuroscientist, Neurological, and so on

Use quotes to search for an exact phrase

"primary prevention of cancer"
(heart or cardiac or cardio*) AND arrest -"American Heart Association"