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Latest Articles

Isolation of Thymic Epithelial Cells and Analysis by Flow Cytometry
Reema Jain, Daniel H.D. Gray

The epithelial cells of the thymus govern the differentiation of hematopoietic precursors into T cells, which are critical for acquired immunity.

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Sampling of Intestinal Microbiota and Targeted Amplification of Bacterial 16S rRNA Genes for Microbial Ecologic Analysis
Maomeng Tong, Jonathan P. Jacobs, Ian H. McHardy, Jonathan Braun

Dysbiosis of host‐associated commensal microbiota is emerging as an important factor in risk and phenotype of immunologic, metabolic, and behavioral diseases.

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T Cell Mediated Autoimmune Glomerular Disease in Mice
Joshua D. Ooi, Poh‐Yi Gan, Dragana Odobasic, Stephen R. Holdsworth, A. Richard Kitching

Many forms of glomerulonephritis are mediated by autoimmunity. While autoantibodies are often pathogenic, cell‐mediated immunity plays an important role in a number of forms of rapidly progressive glomerulonephritis.

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Isolation and Purification of Proteasomes from Primary Cells
Nicholas J. Steers, Kristina K. Peachman, Carl R. Alving, Mangala Rao

Proteasomes play an important role in cell homeostasis and in orchestrating the immune response by systematically degrading foreign proteins and misfolded or damaged host cell proteins.

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Vibrio cholerae: Measuring Natural Transformation Frequency
Samit S. Watve, Eryn E. Bernardy, Brian K. Hammer

Many bacteria can become naturally competent to take up extracellular DNA across their outer and inner membranes by a dedicated competence apparatus.

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Laboratory Maintenance of Acinetobacter baumannii
Anna C. Jacobs, Daniel V. Zurawski

Acinetobacter baumannii has recently drawn great interest in the microbiology research community due to the increase in clinical antibiotic resistance of this organism, and persistence of this bacterial species in the hospital environment.

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Genetic Manipulation of Acinetobacter baumannii
Anna C. Jacobs, Mitchell G. Thompson, Michael Gebhardt, Brendan W. Corey, Suleyman Yildirim, Howard A. Shuman, Daniel V. Zurawski

Acinetobacter baumannii is a Gram‐negative nosocomial pathogen of clinical importance.

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Quantitative Analysis of HSV Gene Expression during Lytic Infection
Anne‐Marie W. Turner, Jesse H. Arbuckle, Thomas M. Kristie

Herpes Simplex Virus (HSV) is a human pathogen that establishes latency and undergoes periodic reactivation, resulting in chronic recurrent lytic infection.

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Analysis of HSV Viral Reactivation in Explants of Sensory Neurons
Jesse H. Arbuckle, Anne‐Marie W. Turner, Thomas M. Kristie

As with all Herpesviruses, Herpes simplex virus (HSV) has both a lytic replication phase and a latency‐reactivation cycle. During lytic replication, there is an ordered cascade of viral gene expression that leads to the synthesis of infectious viral progeny.

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Preparation of Soluble Proteins from Escherichia coli
Paul T. Wingfield

Purification of human IL‐1β is used in this unit as an example of the preparation of a soluble protein from E. coli.

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Folding and Purification of Insoluble (Inclusion Body) Proteins from Escherichia coli
Paul T. Wingfield, Ira Palmer, Shu‐Mei Liang

Heterologous expression of recombinant proteins in E. coli often results in the formation of insoluble and inactive protein aggregates, commonly referred to as inclusion bodies. To obtain the native (i.e.

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Analysis of Protein Composition Using Multidimensional Chromatography and Mass Spectrometry
Andrew J. Link, Michael P. Washburn

Multidimensional liquid chromatography of peptides produced by protease digestion of complex protein mixtures followed by tandem mass spectrometry can be coupled with automated database searching to identify large numbers of proteins in complex samples.

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Proteomic Analysis of Protein Deamidation
Piliang Hao, Siu Kwan Sze

Deamidation of asparagines and glutamines occurs spontaneously in proteins and results in protein degradation.

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Pulse‐Chase Analysis for Studying Protein Synthesis and Maturation
Susanne Fritzsche, Sebastian Springer

Pulse‐chase analysis is a well‐established and highly adaptable tool for studying the life cycle of endogenous proteins, including their synthesis, folding, subunit assembly, intracellular transport, post‐translational processing, and degradation.

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Generation of Mammalian Offspring by Haploid Embryonic Stem Cells Microinjection
Ling Shuai, Wei Li, Haifeng Wan, Xiao‐Yang Zhao, Liu Wang, Qi Zhou

In this unit we introduce the derivation and genetic modification of mouse haploid embryonic stem (ES) cells. We detail how to produce haploid embryos and the subsequent ES derivation and cell culture.

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Cryopreservation of Human Pluripotent Stem Cells in Defined Medium
Weiwei Liu, Guokai Chen

This unit describes a cryopreservation procedure using an enzyme‐free dissociation method to harvest cells and preserve cells in albumin‐free chemically defined E8 medium for human pluripotent stem cells (hPSCs).

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Isolation and Long‐Term Expansion of Functional, Myelinating Oligodendrocyte Progenitor Cells from Neonatal Rat Brain
Bangfu Zhu, Chao Zhao, Fraser I. Young, Robin J.M. Franklin, Bing Song

Oligodendrocytes are the myelinating cells of the central nervous system (CNS).

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Laser‐Assisted Generation of Human Induced Pluripotent Stem Cells
Yang Liu, Dongmei Wu, Donglin Lao, Cory Peterson, Kristi A. Hohenstein Elliott, Evan Y. Snyder

This unit describes a procedure for generating human induced pluripotent stem cells (hiPSCs) using the Laser‐Enabled Analysis and Processing (LEAP®) system, which combines high‐throughput cell imaging with laser‐mediated cell manipulation.

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F3‐Isoprostanes as a Measure of in vivo Oxidative Damage in Caenorhabditis elegans
Thuy T. Nguyen, Michael Aschner

Oxidative stress has been implicated in the development of a wide variety of disease processes, including cardiovascular disease, cancer, and neurodegenerative diseases, as well as progressive and normal aging processes.

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The History and Use of Human Hepatocytes for the Treatment of Liver Diseases: The First 100 Patients
Marc C. Hansel, Roberto Gramignoli, Kristen J. Skvorak, Kenneth Dorko, Fabio Marongiu, William Blake, Julio Davila, Stephen C. Strom

Orthotopic liver transplantation remains the only curative treatment for many end‐stage liver diseases, yet the number of patients receiving liver transplants remains limited by the number of organs available for transplant. There is a need for alternative therapies for liver diseases.

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