• Volume 11, 2021
  • Volume 10, 2020
  • Volume 9, 2019
  • Volume 8, 2018
  • Volume 7, 2017
  • Volume 6, 2016
  • Volume 5, 2015
  • Volume 4, 2014
  • Volume 3, 2013
  • Volume 2, 2012
  • Volume 1, 2011
Image Credit: BioProtoc.3907

Past Issues in 2021

Volume: 11 Issue: 3

Biochemistry

Using 14C-acetate Pulse-chase Labeling to Study Fatty Acid and Glycerolipid Metabolism in Plant Leaves Authors:  Linhui Yu, Chao Zhou, Jilian Fan and Changcheng Xu, date: 02/05/2021, view: 296, Q&A: 0

Lipids metabolism is comprised of networks of reactions occurred in different subcellular compartments. Isotopic labeling is a good way to track the transformations and movements of metabolites without perturbing overall cellular metabolism. Fatty acids, the building blocks of membrane lipids and storage triacylglycerols, are synthesized in

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Purification of Cytosolic Phospholipase A2α C2-domain after Expression in Soluble Form in Escherichia coli Authors:  Yoshinori Hirano, Yong-Guang Gao, Dhirendra K. Simanshu, Daniel J. Stephenson, Ngoc T. Vu, Lucy Malinina, Charles E. Chalfant, Dinshaw J. Patel and Rhoderick E. Brown, date: 02/05/2021, view: 232, Q&A: 0

Previous expression/purification strategies for cytosolic phospholipase A2α C2-domain in Escherichia coli have relied on refolded protein recovered from inclusion bodies and sometimes containing C-terminal Cys139Ala and Cys141Ser substitutions to eliminate potential refolding complications induced by Cys residues. The protocol presented

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Biophysics

Resolving Structural Changes of Photoreceptors in Living Escherichia coli via In-cell Infrared Difference Spectroscopy Authors:  Lukas Goett-Zink, Jessica L. Klocke and Tilman Kottke, date: 02/05/2021, view: 387, Q&A: 0

Several in-cell spectroscopic techniques have been developed recently to investigate the structure and mechanism of proteins in their native environment. Conditions in vivo differ dramatically from those selected for in vitro experiments. Accordingly, the cellular environment can affect the protein mechanism for example by molecular crowding or

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Cancer Biology

Carboxyfluorescein Dye Uptake to Measure Connexin-mediated Hemichannel Activity in Cultured Cells Authors:  Joe A. Potter, Gareth W. Price, Chelsy L. Cliff, Bethany M. Williams, Claire E. Hills and Paul E. Squires, date: 02/05/2021, view: 575, Q&A: 0

Connexins are membrane bound proteins that facilitate direct and local paracrine mediated cell-to-cell communication through their ability to oligomerise into hexameric hemichannels. When neighbouring channels align, they form gap-junctions that provide a direct route for information transfer between cells. In contrast to intact gap junctions,

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Isolation of Murine Primary Aortic Smooth Muscle Cells Authors:  Max Ole Hubert, Juan Rodriguez-Vita, Lena Wiedmann and Andreas Fischer, date: 02/05/2021, view: 244, Q&A: 0

Vascular smooth muscle cells (VSMCs) have been cultured for decades to study the role of these cells in cardiovascular disorders. The most common source of VSMCs is the rat aorta. Here we show the adaptation of this method to isolate and culture mouse aortic VSMCs. The advantage of this method is that there are many more transgenic mouse lines

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Developmental Biology

Induction of Epithelial-mesenchymal Transition in MDCK II Cells Authors:  Agnieszka Pastuła and Richard Lundmark, date: 02/05/2021, view: 284, Q&A: 0

Epithelial-mesenchymal transition (EMT) is a reversible process of epithelial cell transdifferentiation into a mesenchymal cell, that enables initiation of cell migration. EMT plays an important role in embryonic development, tissue repair and cancer metastasis. Better understanding of cellular and molecular events during EMT will not only provide

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Ecdysone Quantification from Whole Body Samples of Drosophila melanogaster Larvae Authors:  Takashi Koyama and Christen K. Mirth, date: 02/05/2021, view: 214, Q&A: 0

Steroid hormones strictly control the timing of sexual maturation and final body size both in vertebrates and invertebrates. In insects, the steroid hormone ecdysone controls the timing of the molts between larval instars as well as the transition to metamorphosis. Growth during the final instar accounts for over 80% of the increase in final mass

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Immunology

A Versatile Protocol to Quantify BCR-mediated Phosphorylation in Human and Murine B Cell Subpopulations Authors:  Jasper Rip, Rudi W. Hendriks and Odilia B. J. Corneth, date: 02/05/2021, view: 301, Q&A: 0

Signal transduction is the process by which molecular signals are transmitted from the cell surface to its interior, resulting in functional changes inside the cell. B cell receptor (BCR) signaling is of crucial importance for B cells, as it regulates their differentiation, selection, survival, cellular activation and proliferation. Upon BCR

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In vitro STING Activation with the cGAMP-STINGΔTM Signaling Complex Authors:  Yanpu He, Celestine Hong, Darrell J. Irvine, Jiahe Li and Paula T. Hammond, date: 02/05/2021, view: 295, Q&A: 0
Activating the STING (stimulator of interferon genes) signaling pathway via administration of STING agonist cyclic GMP-AMP (cGAMP) has shown great promise in cancer immunotherapy. While state-of-the-art approaches have predominantly focused on the encapsulation of cGAMP into liposomes or polymersomes for cellular delivery, we discovered that the ...
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Neuroscience

Relative Quantification of NaV1.1 Protein in Mouse Brains Using a Meso Scale Discovery-Electrochemiluminescence (MSD-ECL) Method Authors:  Zhou Han, Anne Christiansen, Meena and Gene Liau, date: 02/05/2021, view: 225, Q&A: 0

Densitometric analysis is often used to quantify NaV1.1 protein on immunoblots, although the sensitivity and dilution linearity of the method are usually poor. Here we present a protocol for quantification of NaV1.1 in mouse brain tissues using a Meso Scale Discovery-Electrochemiluminescence (MSD-ECL) method. MSD-ECL is based on ELISA

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Immuno-electrophysiology on Neuromuscular Junctions of Drosophila Third Instar Larva Authors:  Raffaella Klima, Giulia Romano, Monsurat Gbadamosi, Aram Megighian and Fabian Feiguin, date: 02/05/2021, view: 215, Q&A: 0

Alterations in synaptic transmission are critical early events in neuromuscular disorders. However, reliable methodologies to analyze the functional organization of the neuromuscular synapses are still needed. This manuscript provides a detailed protocol to analyze the molecular assembly of the neuromuscular synapses through

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Plant Science

Histochemical Staining of Suberin in Plant Roots Authors:  Peter Marhavý and Shahid Siddique, date: 02/05/2021, view: 423, Q&A: 0

Histological stains are useful tools for characterizing cell shape, arrangement and the material they are made from. Stains can be used individually or simultaneously to mark different cell structures or polymers within the same cells, and to visualize them in different colors. Histological stains can be combined with genetically-encoded

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Measuring Extracellular Proton and Anionic Fluxes in Arabidopsis Pollen Tubes Authors:  Maria Teresa Portes and José A. Feijó, date: 02/05/2021, view: 216, Q&A: 0

The ion-selective vibrating probe has been used to detect and quantify the magnitude and direction of transmembrane fluxes of several ions in a wide range of biological systems. Inherently non-invasive, vibrating probes have been essential to access relevant electrophysiological parameters related to apical growth and morphogenesis in pollen

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Plant ARGONAUTE Protein Immunopurification for Pathogen Cross Kingdom Small RNA Analysis Authors:  Florian Dunker, Bernhard Lederer and Arne Weiberg, date: 02/05/2021, view: 355, Q&A: 0

Over the last decade, it has been noticed that microbial pathogens and pests deliver small RNA (sRNA) effectors into their host plants to manipulate plant physiology and immunity for infection, known as cross kingdom RNA interference. In this process, fungal and oomycete parasite sRNAs hijack the plant ARGONAUTE (AGO)/RNA-induced silencing complex

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A Pipeline for Non-model Organisms for de novo Transcriptome Assembly, Annotation, and Gene Ontology Analysis Using Open Tools: Case Study with Scots Pine Authors:  Gustavo T. Duarte, Polina Yu. Volkova and Stanislav A. Geras’kin, date: 02/05/2021, view: 475, Q&A: 0

RNA sequencing (RNA-seq) has opened up the possibility of studying virtually any organism at the whole transcriptome level. Nevertheless, the absence of a sequenced and accurately annotated reference genome may be an obstacle for applying this technique to non-model organisms, especially for those with a complex genome. While de novo transcriptome

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Stem Cell

Rapid and Simplified Induction of Neural Stem/Progenitor Cells (NSCs/NPCs) and Neurons from Human Induced Pluripotent Stem Cells (hiPSCs) Authors:  Ryutaro Kajihara, Tadahiro Numakawa and Takumi Era, date: 02/05/2021, view: 271, Q&A: 0

Human induced pluripotent stem cells (iPSCs) and their progeny displaying tissue-specific characteristics have paved the way for regenerative medicine and research in various fields such as the elucidation of the pathological mechanism of diseases and the discovery of drug candidates. iPSC-derived neurons are particularly valuable as it is

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