JoVE Immunology and Infection
1School of Chemistry, Food and Pharmacy, The University of Reading, 2Department of Nutritional Sciences, The University of Reading
A progressive colonization procedure is described to further assess its impact on the host hepatic metabolism. Colonization is monitored non invasively by evaluating the urinary excretion of microbial co-metabolites by NMR-based metabolic profiling while hepatic metabolism is assessed by High Resolution Magic Angle Spinning (HR MAS) NMR profiling of intact biopsy.
Published December 15, 2011. Keywords: Immunology, Germ-free animal, colonization, NMR, HR MAS NMR, metabonomics
1School of Molecular Biosciences, Washington State University, 2Center for Reproductive Biology, Washington State University
Caenorhabditis elegans is a useful model to explore the functions of polyunsaturated fatty acids in development and physiology. This protocol describes an efficient method of supplementing the C. elegans diet with polyunsaturated fatty acids.
Published November 29, 2013. Keywords: Biochemistry, Caenorhabditis elegans, C. elegans, Nutrition Therapy, genetics (animal and plant), Polyunsaturated fatty acids, omega-6, omega-3, dietary fat, dihomo-gamma-linolenic acid, germ cells
1Department of Craniofacial Development and Stem Cell Biology, and Department of Orthodontics, Dental Institute, Guy's Hospital, UK, King's College London, 2Department of Pediatric Dentistry and Orthodontics, College of Dentistry, King Saud University, Kingdom of Saudi Arabia
Here we detail a method to culture tooth germs in mandible slices using a tissue chopper. This method allows unique access to the tooth during development, providing excellent opportunity for manipulation and lineage tracing, not available using more traditional culture methods.
Published November 13, 2013. Keywords: Anatomy, Tooth, Culture Techniques, Embryo Culture Techniques, Organ Culture Techniques, Developmental Biology, animal biology, animal models, Tooth germ, live slice, development, tissue chopper, lineage tracing, molar, incisor, gland
1Department of Surgery, Weill Cornell Medical College
A simple method to derive and maintain spermatogonial stem and progenitor cell lines from adult mice is presented here. The method utilizes feeder cells originating from the somatic cell compartment of the adult mouse testis. This technique is applicable to common mouse strains, including transgenic, knock-out, and knock-in mice.
Published February 25, 2013. Keywords: Stem Cell Biology, Molecular Biology, Cellular Biology, Medicine, Genetics, Developmental Biology, Anatomy, Surgery, Spermatogonial Stem cells, Stem cells, feeder cells, germ cells, testis, cell culture, microenvironment, stem cell niche, progenitor cells, mice, transgenic mice, animal model
1Dominick P. Purpura Department of Neuroscience, Albert Einstein College of Medicine, Yeshiva University
We provide a practical guide for delivering tracers in vivo and use the spinocerebellar pathway as a model system to demonstrate essential steps for successful neuronal circuit analysis in mice. We describe in detail our versatile tracing protocol that exploits wheat germ agglutinin (WGA) conjugated to Alexa fluorophores.
Published November 14, 2011. Keywords: Neuroscience, neuronal projections, topography, circuits, connectivity, fluorescent tracers, mice
1Materials Science and Engineering, CSIRO
We describe the use of the monoclonal antibodies TG30 (CD9) and GCTM-2 for the combined detection of cell surface antigens via fluorescence activated cell sorting (FACS) for the identification and enrichment of live human embryonic stem cells (hESC) using positive selection and also the use of negative selection to purge hESCs from a mixed cell population.
Published December 6, 2013. Keywords: Stem Cell Biology, Stem cells, cell surface antigens, antibodies, FACS, purging stem cells, differentiation, pluripotency, teratoma, human embryonic stem cells (hESC)
1Department of Cellular and Molecular Medicine, University of California, San Diego, 2Biosecurity and Public Health, Los Alamos National Laboratory
Unfixed frozen tissue samples embedded in Optimal Cutting Temperature medium (OCT) can be used to study natural distribution and glycosylation of secreted mucus. In this approach tissue processing is minimal and the natural presentation of glycolipids, mucins and glycan-epitopes is preserved. Tissue sections can be analyzed by immunohistochemistry using fluorescence or chromogenic detection.
Published September 21, 2012. Keywords: Medicine, Cellular Biology, Molecular Biology, Immunology, Biomedical Engineering, mucus, lectins, OCT, imaging, sialic acids, glycosylation
1San Diego Regenerative Medicine Institute, 2Xcelthera, 3Department of Neurosurgery, Harvard Medical School, 4Division of SCI Research, VA Boston Healthcare System, 5Program in Stem Cell & Regenerative Biology, Sanford-Burnham Medical Research Institute, 6La Jolla IVF
We have established a protocol for induction of neuroblasts direct from pluripotent human embryonic stem cells maintained under defined conditions with small molecules, which enables derivation of a large supply of human neuronal progenitors and neuronal cell types in the developing CNS for neural repair.
Published October 28, 2011. Keywords: Neuroscience, stem cell, human embryonic stem cell, human, neuronal progenitor, neuron, human pluripotent cell, neuronal differentiation, small molecule induction, cell culture, cell therapy
1Biology Department, California State University
In model organisms, transgenesis can manipulate gene functions while RNAi can knockdown specific mRNA transcripts 1-2. This protocol aims to illustrate the techniques needed to introduce stably transmitted DNA and transient double stranded RNA into the necromenic nematode Pristionchus pacificus for studies in evolutionary, developmental, and behavioral biology.
Published October 16, 2011. Keywords: Developmental Biology, RNA interference, Pristionchus pacificus, microinjection, transgenesis, Caenorhabditis elegans, developmental biology, behavior, gene expression
1Department of Biological Sciences, Ohio University
"Freeze-cracking," a method for exposing the inner tissues of the nematode C. elegans to antibodies for protein localization, is demonstrated.
Published October 14, 2013. Keywords: Molecular Biology, Caenorhabditis elegans (C. elegans), Fluorescent Antibody Technique, nematode, labeling, localization, in situ