Volume: 7 Issue: 21 —BIO-PROTOCOL

Biochemistry

Monitoring the Targeting of Cathepsin D to the Lysosome by Metabolic Labeling and Pulse-chase Analysis Authors: Lucas A. Tavares and Luis L. P. daSilva, date: 11/05/2017, view: 4452, Q&A: 0

Mannose 6-phosphate receptors function can be studied in living cells by investigating alterations in processing and secretion of their ligand Cathepsin D. The assay described here is well established in the literature and comprises the metabolic labeling of newly synthesized proteins with [³⁵S] methionine-cysteine in HeLa cells to ...

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Preparation of the Partially Methylated Alditol Acetates Derived from CS Tetrasaccharides Containing Galactose for the Gas Chromatography/Mass Spectrometry Analysis Authors: Kyohei Higashi and Toshihiko Toida, date: 11/05/2017, view: 5652, Q&A: 0

Chondroitin sulfate (CS), a member of the glycosaminoglycan (GAG) family of carbohydrates, is composed of linear, sulfated repeating disaccharide sequences of N-acetyl-_D-galactosamine (GalNAc) and glucuronic acid (GlcA). Recently, a keratan sulfate (KS) disaccharide [GlcNAc6S(β1-3)Galactose(β1-]-branched CS-E was identified ...

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Cancer Biology

In vitro NLK Kinase Assay Authors: Sungho Moon, Jiyoung Kim and Eek‐hoon Jho, date: 11/05/2017, view: 6307, Q&A: 0

This protocol provides step by step instructions to perform an in vitro kinase assay for nemo-like kinase. In addition, this protocol also describes an efficient method using mild lysis buffer for expression and purification of Glutathione S-transferase (GST) fusion proteins.

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Xenograft Mouse Model of Human Uveal Melanoma Authors: Yao Chen, Xiao Liu, Ling Gao and Yongqing Liu, date: 11/05/2017, view: 4627, Q&A: 0

Uveal melanoma (UM) is a malignant intraocular tumor in adults. Metastasis develops in almost half of the patients and over 90% of the metastases are in the liver. With the advances in molecular targeting therapy for melanoma, a proper metastasis animal model is of increasing importance for testing the accuracy and effectiveness of systemic ...

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Cell Biology

A Bioreactor Method to Generate High-titer, Genetically Stable, Clinical-isolate Human Cytomegalovirus Authors: Victoria R. Saykally, Luke I. Rast, Jeff Sasaki, Seung-Yong Jung, Cynthia Bolovan-Fritts and Leor S. Weinberger, date: 11/05/2017, view: 5066, Q&A: 0

Human cytomegalovirus (HCMV) infection is a major cause of morbidity and mortality in transplant patients and a leading cause of congenital birth defects (Saint Louis, 2016). Vaccination and therapeutic studies often require scalable cell culture production of wild type virus, represented by clinical isolates. Obtaining sufficient stocks of ...

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Histochemical Staining of Collagen and Identification of Its Subtypes by Picrosirius Red Dye in Mouse Reproductive Tissues Authors: Smita Bhutda, Manalee V Surve, Anjali Anil, Kshama Kamath, Neha Singh, Deepak Modi and Anirban Banerjee , date: 11/05/2017, view: 15437, Q&A: 1

Collagen is one of the foremost components of tissue extracellular matrix (ECM). It provides strength, elasticity and architecture to the tissue enabling it to bear the wear and tear from external factors like physical stress as well as internal stress factors like inflammation or other pathological conditions. During normal pregnancy or pregnancy ...

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Nematode Epicuticle Visualisation by PeakForce Tapping Atomic Force Microscopy Authors: Farida Akhatova, Gölnur Fakhrullina, Elvira Gayazova and Rawil Fakhrullin, date: 11/05/2017, view: 4647, Q&A: 0

The free-living soil nematode Caenorhabditis elegans has become an iconic experimental model animal in biology. This transparent animal can be easily imaged using optical microscopy to visualise its organs, tissues, single cells and subcellular events. The epicuticle of C. elegans nematodes has been studied at nanoscale using ...

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Immunogold Localization of Molecular Constituents Associated with Basal Bodies, Flagella, and Extracellular Matrices in Male Gametes of Land Plants Authors: Renee A. Lopez, Katayoun Mansouri, Jason S. Henry, Nicholas D. Flowers, Kevin C. Vaughn and Karen Sue Renzaglia, date: 11/05/2017, view: 4215, Q&A: 0

Male gametes (spermatozoids) are the only motile cells produced during the life cycle of land plants. While absent from flowering and most cone-bearing plants, motile cells are found in less derived taxa, including bryophytes (mosses, liverworts and hornworts), pteridophytes (lycophytes and ferns) and some seed plants (Ginkgo and cycads). ...

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Immunology

Proximal Ligation Assay (PLA) on Lung Tissue and Cultured Macrophages to Demonstrate Protein-protein Interaction Authors: Roberto Mendez and Santanu Banerjee, date: 11/05/2017, view: 6049, Q&A: 0

In this protocol, we describe proximal ligation assay (PLA), an antibody-based detection method for protein-protein interaction. This method relies on specific binding of individual primary antibodies to the two putative interacting proteins. The primary antibodies need to have different hosts. The secondary antibodies against the two hosts have ...

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Microbiology

Isolation of Rice Stripe Virus Preparation from Viruliferous Small Brown Planthoppers and Mechanic Inoculation on Rice Authors: Wan Zhao, Le Kang and Feng Cui, date: 11/05/2017, view: 3586, Q&A: 0

Tenuiviruses can infect the plants of the family Poaceae, and cause serious loss of crops, particularly rice and maize, in South-Eastern Asian countries. Tenuiviruses usually depend on insect vectors for their transmission and cannot be transmitted between plants through wounds or abrasions. Rice stripe virus (RSV), a typical member of ...

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Neuroscience

Assessment of Aversion of Acute Pain Stimulus through Conditioned Place Aversion Authors: Louise Urien, Qiaosheng Zhang, Erik Martinez, Haocheng Zhou, Nicole Desrosier, Jahrane Dale and Jing Wang, date: 11/05/2017, view: 4331, Q&A: 0

Pain is a complex experience. The aversive component of pain has been assessed through conditioned place aversion in rodents. However, this behavioral test does not allow the evaluation of the aversion of an acute pain stimulus. In Zhang et al. (2017), we provide an updated version of a Conditioned Place Aversion paradigm to address this ...

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Sensitive Estimation of Flavor Preferences in STFP Using Cumulative Time Profiles Authors: Aditya Singh and J. Balaji, date: 11/05/2017, view: 3834, Q&A: 0

Social transmission of food preference (STFP) is observed among rodents between a demonstrator and a naïve hungry observer. During social interaction, hungry observer receives information about safety of the food consumed by the demonstrator. This task has been implemented to develop a single trial non-aversive learning task in order to test ...

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Isolation, Culture and Differentiation of Adult Hippocampal Precursor Cells Authors: Stefanie N. Bernas, Odette Leiter, Tara L. Walker and Gerd Kempermann, date: 11/05/2017, view: 9447, Q&A: 0

There are two neurogenic niches in the adult mammalian brain: the subventricular zone of the lateral ventricle and the subgranular zone of the hippocampal dentate gyrus. Cells from these areas can be isolated and maintained in vitro, using two different culture systems to assess their potential regarding proliferation and differentiation ...

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Stem Cell

Isolation of Primary Human Skeletal Muscle Cells Authors: Janelle M. Spinazzola and Emanuela Gussoni, date: 11/05/2017, view: 4994, Q&A: 0

Primary myoblast culture is a valuable tool in research of muscle disease, pathophysiology, and pharmacology. This protocol describes techniques for dissociation of cells from human skeletal muscle biopsies and enrichment for a highly myogenic population by fluorescence-activated cell sorting (FACS). We also describe methods for assessing ...

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Past Issues in 2017

Volume: 7 Issue: 21

Biochemistry

Cancer Biology

Cell Biology

Immunology

Microbiology

Neuroscience

Stem Cell