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Nonhuman Primate Models in Trophic Factor Research

Nonhuman primates are essential experimental models for research dealing with trophic factors, particularly regarding the effects of these factors on higher cognitive functions, including memory. These animals have several advantages over rodents. First, these species are o ...

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Experimental Systems to Study Neurotrophic Factor Effects on Rat Brain Cells

Nerve growth factor (NGF) was discovered and the concept of neurotrophic factors as retrograde messengers was developed in studies on the peripheral nervous system (PNS). Initial descriptions of biological effects of newly discovered grow factors often are limited to PNS neurons, whe ...

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Depletion of Microglia from Primary Cellular Cultures

Primary cultures are an important in vitro tool to study cellular processes and interactions. These cultures are complex systems, composed of many cell types, including neurons, astrocytes, oligodendrocytes, microglia, NG2 cells, and endothelial cells. For some studies it is necess ...

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Isolation and Culture of Adult Human Microglia Within Mixed Glial Cultures for Functional Experimentation and High-Content Analysis

Microglia are thought to be involved in diseases of the adult human brain as well as normal aging processes. While neonatal and rodent microglia are often used in studies investigating microglial function, there are important differences between rodent microglia and their adult human co ...

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Isolation of Murine Postnatal Brain Microglia for Phenotypic Characterization Using Magnetic Cell Separation Technology

To shorten the time between brain harvesting and microglia isolation, and characterization, we utilized the MACS� neural dissociation kit followed by OctoMACS� CD11b magnetic bead isolation technique to positively select for brain microglia expressing the pan-microglial mar ...

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Preparation of Primary Microglia Cultures from Postnatal Mouse and Rat Brains

Microglia are the inflammatory cells of the brain and are activated in neuropathological conditions. To study the biology of microglia, these cells can be isolated from the brain and analyzed in terms of pro- and anti-inflammatory cytokine production, involvement of intracellular sig ...

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Cell Culturing of Human and Murine Microglia Cell Lines

Despite the fact that microglia cells were first described almost a century ago, microglia-derived immortalized cell lines have only been established in the last two decades. One should be aware of their limitations but also of their advantages. Cell lines offer a potentially powerful tool to ...

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Microglia Isolation from Adult Mouse Brain

Although microglia isolation from embryonic or postnatal mouse brain is possible using a number of different protocols, microglia isolation from adult brain is more challenging and often results in low yields. Here, we describe a protocol to isolate intact microglia from adult mouse bra ...

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Isolating, Culturing, and Polarizing Primary Human Adult and Fetal Microglia

Microglia are an important component of the innate immune system within the central nervous system (CNS). Isolation and in vitro culturing of microglia can provide insight towards the basic biology of these cells as well as their interactions with neurons, astrocytes, and oligodendrocy ...

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Studying M1 and M2 States in Adult Microglia

Microglial cell function receives increasing interest. To date, the majority of experiments are performed by using immortalized microglia-like cells or primary microglia prepared from pre- or postnatal rodent brain. As those may not adequately reflect the microglial biology in the ...

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Patch Clamp Protocols to Study Ion Channel Activity in Microglia

Microglia express a variety of ion channels, which can be distinguished based on their ion selectivity into K+, H+, Na+, Ca2+, nonselective cation, and Cl− channels. With respect to their activation mode, voltage-, Ca2+-, calcium release-, G protein-, swelling-, and stretch-activated ion channels ...

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Fluorescence Imaging of Intracellular Ca2+, Na+, and H+ in Cultured Microglia

The behavior of microglial cells involves the activity of a variety of ion channels and ion transporters, which are implicated in the regulation of ion concentrations, membrane potential, and cell volume of microglia. Fluorescence imaging has been proven to be an elegant method to study ion co ...

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Quantification of Microglial Phagocytosis by a Flow Cytometer-Based Assay

Microglia represent the largest population of phagocytes in the CNS and have a principal role in immune defense and inflammatory responses in the CNS. Their phagocytic activity can be studied by a variety of techniques, including a flow cytometry-based approach utilizing polystyrene l ...

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Quantification of Microglial Proliferation and Apoptosis by Flow Cytometry

Microglia are innate immune cells that survey the central nervous system (CNS) and respond almost immediately to any disturbance in CNS homeostasis. They are derived from primitive yolk sac myeloid progenitors and in the mouse colonize the CNS during fetal development. As a population, mic ...

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Quantification of Active Caspase-3 and Active Caspase-8 in Microglia Cells

During microglia activation the levels of active caspase-3, caspase-7 and caspase-8 are increased, which leads to the transcription of proinflammatory cytokines and factors. As such, the induction of caspase activity in microglia can be used as a marker for activation. The use of sensitive a ...

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Understanding MicrogliaNeuron Cross Talk: Relevance of the MicrogliaNeuron Cocultures

Microglia–neuron interaction is a complex process involving a plethora of ligands and receptors. The outcome of this intricate process will depend on the prevailing signals (i.e., whether the microglial cells will produce pro-inflammatory cytokines and/or phagocyte a dying neuron ...

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Immunohistochemical Detection of Microglia

Immunohistochemistry (IHC) is a technique that allows the localization of antigens or proteins in tissue sections using the high specificity and affinity of antibodies to recognize molecules and join them. The commercial offer and the standardization of protocols make this techniq ...

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Tomato Lectin Histochemistry for Microglial Visualization

The use of different lectins for the study of microglial cells in the central nervous system (CNS) is a valuable tool that has been extensively used in the last years for the selective staining of this glial cell population, not only in normal physiological conditions, but also in a wide range of patholo ...

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Microglia Detection by Enzymatic Histochemistry

Visualization of microglia by means of histochemistry has been for years a reliable method to demonstrate this population of cells in the central nervous system (CNS). Wide range of data on microglia has been published using lectin and enzymatic histochemistry. While at present, in most labo ...

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Preparation of Rodent Primary Cultures for NeuronGlia, Mixed Glia, Enriched Microglia, and Reconstituted Cultures with Microglia

Microglia, neurons, and macroglia (astrocytes and oligodendrocytes) are the major cell types in the central nervous system. In the past decades, primary microglia-enriched cultures have been widely used to study the biological functions of microglia in vitro. In order to study the inter ...

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