• Volume 12, 2022
  • 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.4360

Past Issues in 2022

Volume: 12 Issue: 6

Biochemistry

Analysis of Heterocyst and Akinete Specific Glycolipids in Cyanobacteria Using Thin-layer Chromatography Authors:  Ritu Garg, Rebeca Perez and Iris Maldener, date: 03/20/2022, view: 509, Q&A: 0

Several filamentous cyanobacteria like Nostoc differentiate specialized cells in response to changes in environmental factors, such as low light or nutrient starvation. These specialized cells are termed heterocysts and akinetes. Under conditions of nitrogen limitation, nitrogen-fixing heterocysts form in a semi-regular pattern and provide the

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Novel Protein Production Method Combining Native Expression in Human Cells with an Intein-based Affinity Purification and Self-cleavable Tag Authors:  Peter J. Carman and Roberto Dominguez, date: 03/20/2022, view: 448, Q&A: 0

The human proteins used in most biochemical studies are commonly obtained using bacterial expression. Owing to its relative simplicity and low cost, this approach has been extremely successful, but is inadequate for many proteins that require the mammalian folding machinery and posttranslational modifications (PTMs) for function. Moreover, the

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Biological Engineering

A Fluorescence-based Assay for Measuring Phospholipid Scramblase Activity in Giant Unilamellar Vesicles Authors:  Patricia P. M. Mathiassen and Thomas Günther Pomorski, date: 03/20/2022, view: 525, Q&A: 0

Transbilayer movement of phospholipids in biological membranes is mediated by a diverse set of lipid transporters. Among them are scramblases that facilitate rapid bi-directional movement of lipids without metabolic energy input. In this protocol, we describe the incorporation of phospholipid scramblases into giant unilamellar vesicles (GUVs)

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Biophysics

Electrophysiological Measurements of Isolated Blood Vessels Authors:  Samuel A Molina, Daniela Maier-Begandt, Brant E Isakson and Michael Koval, date: 03/20/2022, view: 415, Q&A: 0

The lumen of blood vessels is covered by endothelial cells, which regulate their permeability to ions and solutes. Endothelial permeability depends on the vascular bed and cell phenotype, and is influenced by different disease states. Most characterization of endothelial permeability has been carried out using isolated cells in culture. While

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Cancer Biology

Small Molecule Screening of Primary Human Acute Myeloid Leukemia Using Co-culture and Multiplexed FACS Analysis Authors:  Aurelie Baudet, Simon Hultmark, Fredrik Ek and Mattias Magnusson, date: 03/20/2022, view: 439, Q&A: 0

Ex vivo culture of primary acute myeloid leukemia (AML) cells is notoriously difficult due to spontaneous differentiation and cell death, which hinders mechanistic and translational studies. To overcome this bottleneck, we have implemented a co-culture system, where the OP9-M2 stromal cells support the growth, but most notably limit the

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Single-cell Damagenome Profiling by Linear Copying and Splitting based Whole Genome Amplification (LCS-WGA) Authors:  Yichi Niu, Qiangyuan Zhu and Chenghang Zong, date: 03/20/2022, view: 480, Q&A: 0

Spontaneous DNA damage frequently occurs on the human genome, and it could alter gene expression by inducing mutagenesis or epigenetic changes. Therefore, it is highly desired to profile DNA damage distribution on the human genome and identify the genes that are prone to DNA damage. Here, we present a novel single-cell whole-genome amplification

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Developmental Biology

TUNEL Labeling to Detect Double-stranded DNA Breaks in Caenorhabditis elegans Gonads Authors:  Peter A. Kropp, Kyle Rhodehouse and Andy Golden, date: 03/20/2022, view: 405, Q&A: 0

Analysis of DNA double strand breaks (DSBs) is important for understanding dyshomeostasis within the nucleus, impaired DNA repair mechanisms, and cell death. In the C. elegans germline, DSBs are important indicators of all three above-mentioned conditions. Although multiple methods exist to assess apoptosis in the germline of C. elegans, direct

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3D-Structured Illumination Microscopy of Centrosomes in Human Cell Lines Authors:  Kari-Anne M. Frikstad, Kay O. Schink, Sania Gilani, Lotte B. Pedersen and Sebastian Patzke, date: 03/20/2022, view: 699, Q&A: 0

The centrosome is the main microtubule-organizing center of animal cells, and is composed of two barrel-shaped microtubule-based centrioles embedded in protein dense pericentriolar material. Compositional and architectural re-organization of the centrosome drives its duplication, and enables its microtubule-organizing activity and capability to

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Measurement of Contractile Ring Tension Using Two-photon Laser Ablation during Drosophila Cellularization Authors:  Swati Sharma and Richa Rikhy, date: 03/20/2022, view: 372, Q&A: 0

Cytokinesis occurs at the final step of cell division and leads to the separation of daughter cells. It requires assembly and constriction of the actomyosin contractile ring. The phases of assembly and constriction of the contractile ring show an increase in tension in the actomyosin complex. The measurement of tension in the contractile ring is

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Immunology

CD8 T Cell Virus Inhibition Assay Protocol Authors:  Yinyan Xu, Ann Marie Weideman, Maria Abad-Fernandez, Katie R. Mollan, Sallay Kallon, Shahryar Samir, Joanna A. Warren, Genevieve Clutton, Nadia Roan, Adaora A. Adimora, Nancie Archin, JoAnn Kuruc, Cindy Gay, Michael G. Hudgens and Nilu Goonetilleke, date: 03/20/2022, view: 446, Q&A: 0

The human immunodeficiency virus (HIV)-1 viral inhibition assay (VIA) measures CD8+ T cell-mediated inhibition of HIV replication in CD4+ T cells and is increasingly used for clinical testing of HIV vaccines and immunotherapies. Different VIAs that differ in length of CD8:CD4 T cell culture periods (6–13 days), purity of CD4 cultures

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Differentiation of Human Induced Pluripotent Stem Cell into Macrophages Authors:  Harriet Douthwaite, Aitor Bermejo Arteagabeitia and Subhankar Mukhopadhyay, date: 03/20/2022, view: 823, Q&A: 0

As a model to interrogate human macrophage biology, macrophages differentiated from human induced pluripotent stem cells (hiPSCs) transcend other existing models by circumventing the variability seen in human monocyte-derived macrophages, whilst epitomizing macrophage phenotypic and functional characteristics over those offered by macrophage-like

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Microbiology

Phytophthora sojae Transformation Based on the CRISPR/Cas9 System Authors:  Jingting Cao, Min Qiu, Wenwu Ye and Yuanchao Wang, date: 03/20/2022, view: 518, Q&A: 0

Phytophthora sojae is a model species for the study of plant pathogenic oomycetes. The initial research on gene function using Phytophthora was mainly based on gene silencing technology. Recently, the CRISPR/Cas9-mediated genome editing technology was successfully established in P. sojae and widely used in oomycetes. In this protocol, we describe

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Generation, Maintenance and HBV Infection of Human Liver Organoids Authors:  Shahla Romal, Tanvir Hossain and Tokameh Mahmoudi, date: 03/20/2022, view: 896, Q&A: 0

Hepatitis B virus (HBV) infection represents a major public health problem infecting approximately 400 million people worldwide. Despite the availability of a preventive vaccine and anti-viral therapies, chronic HBV infection remains a major health issue because it increases the risk of developing liver cirrhosis and hepatocellular carcinoma

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Tracking the Reversed Oxidative Tricarboxylic Acid Cycle in Bacteria Authors:  Lydia Steffens, Eugenio Pettinato, Thomas M. Steiner, Wolfgang Eisenreich and Ivan A. Berg, date: 03/20/2022, view: 785, Q&A: 1

Different pathways for autotrophic CO2 fixation can be recognized by the presence of genes for their specific key enzymes. On this basis, (meta)genomic, (meta)transcriptomic, or (meta)proteomic analysis enables the identification of the role of an organism or a distinct pathway in primary production. However, the recently discovered variant of the

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Three-dimensional Models of the Nasopharynx for the Study of Epstein-Barr Virus Infection Authors:  Phillip Ziegler, Alex S. Reznik, Shweta P. Kitchloo, Eric Wang, Stella E. Lee, Anthony Green, Michael M. Myerburg, Clare E. Sample and Kathy Ho Yen Shair, date: 03/20/2022, view: 493, Q&A: 0

The ubiquitous and cancer-associated Epstein-Barr virus (EBV) is associated with nearly all cases of nasopharyngeal carcinoma (NPC). Nasopharyngeal tissue is comprised of both pseudostratified and stratified epithelium, which are modeled in three-dimensional (3-D) cell culture. The cellular origin of EBV-associated NPC is as yet unknown, but both

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Plant Science

Activity-based Protein Profiling of Serine Hydrolase Superfamily Enzymes Authors:  Achintya Kumar Dolui, Mahadev Latha and Panneerselvam Vijayaraj, date: 03/20/2022, view: 485, Q&A: 0

Activity-based protein profiling (ABPP) is a chemoproteomics platform to assess the functional state of enzymes in complex biological systems. Over the two decades, ABPP has emerged from a gel-based to gel-free platform, for in-depth proteome analysis with enhanced resolution, sensitivity for target detection, and discovery of small molecule

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