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Immunofluorescent visualization of mouse interneuron subtypes.

F1000Research | 2014

The activity of excitatory neurons is controlled by a highly diverse population of inhibitory interneurons. These cells show a high level of physiological, morphological and neurochemical heterogeneity, and play highly specific roles in neuronal circuits. In the mammalian hippocampus, these are divided into 21 different subtypes of GABAergic interneurons based on their expression of different markers, morphology and their electrophysiological properties. Ideally, all can be marked using an antibody directed against the inhibitory neurotransmitter GABA, but parvalbumin, calbindin, somatostatin, and calretinin are also commonly used as markers to narrow down the specific interneuron subtype. Here, we describe a journey to find the necessary immunological reagents for studying GABAergic interneurons of the mouse hippocampus. Based on web searches there are several hundreds of different antibodies on the market directed against these four markers. Searches in the literature databases allowed us to narrow it down to a subset of antibodies most commonly used in publications. However, in our hands the most cited ones did not work for immunofluorescence stainings of formaldehyde fixed tissue sections and cultured hippocampal neurons, and we had to immunostain our way through thirteen different commercial antibodies before finally finding a suitable antibody for each of the four markers. The antibodies were evaluated based on signal-to-noise ratios as well as if positive cells were found in layers of the hippocampus where they have previously been described. Additionally, the antibodies were also tested on sections from mouse spinal cord with similar criteria for specificity of the antibodies. Using the antibodies with a high rating on pAbmAbs, an antibody review database, stainings with high signal-to-noise ratios and location of the immunostained cells in accordance with the literature could be obtained, making these antibodies suitable choices for studying the GABAergic system.

Pubmed ID: 25469233 RIS Download

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This is a list of tools and resources that we have found mentioned in this publication.


Abbott Diagnostics (tool)

RRID:SCR_008392

Company offering a broad range of instrument systems and diagnostic tests for hospitals, reference labs, blood banks, physician offices and clinics to aid in the diagnosis of a range of serious health issues such as infectious diseases, cancer, and diabetes, as well as monitor other important indicators of health.

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Dako (tool)

RRID:SCR_013530

An Antibody supplier; Dako was purchased by Agilent in 2012 and several years later the websites began to reflect the Dako products as part of the Agilent catalog.

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Synaptic Systems (tool)

RRID:SCR_013612

An Antibody supplier

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Anti-Somatostatin, N-Terminal antibody produced in rabbit (antibody)

RRID:AB_10747468

This polyclonal targets Somatostatin N-Terminal antibody produced in rabbit

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Anti-Somatostatin Antibody, clone YC7 (antibody)

RRID:AB_2255365

This monoclonal targets Somatostatin

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Anti-Parvalbumin (antibody)

RRID:AB_838238

This polyclonal targets Parvalbumin

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Anti-Parvalbumin (antibody)

RRID:AB_2156476

This polyclonal targets Parvalbumin

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Calretinin (CR) Antibody (antibody)

RRID:AB_10000320

This monoclonal targets Calretinin (CR)

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Anti-Calretinin (Sheep) Antibody - 200-601-D13 (antibody)

RRID:AB_11183443

This unknown targets Calretinin Antibody

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Anti-Calretinin (antibody)

RRID:AB_2068506

This polyclonal targets Calretinin

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Calbindin D28k (antibody)

RRID:AB_2314070

This unknown targets

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Monoclonal Anti-Calbindin-D-28K antibody produced in mouse (antibody)

RRID:AB_476894

This monoclonal targets Calbindin-D-28K antibody produced in mouse

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Anti-Calbindin D-28K (antibody)

RRID:AB_2068336

This polyclonal targets Calbindin D-28K

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ZEISS ZEN Microscopy Software (software resource)

RRID:SCR_013672

User interface software for Carl Zeiss light microscopy imaging systems. ZEN is the universal user interface you will see on every imaging system from ZEISS. After selecting fluorophore, ZEN applies the necessary settings to collect and organize data.

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