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Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the delivery of a neuronal tracer using the Nanoject II. The surgery uses a stereotaxic system to target specific brain coordinates in the mouse. Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Injection of Viral Tracers by Nanoject. protocols.io dx.doi.org/10.17504/protocols.io.bgpujvnw Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used to prepare Artificial Cerebrospinal Fluid VII (ACSF.VII). ACSF.VII is used for applications including fresh human brain tissue transportation and as a bathing solution during fresh human brain slicing.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U01MH114812. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Artificial Cerebrospinal Fluid VII (ACSF.VII). protocols.io dx.doi.org/10.17504/protocols.io.beznjf5e Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the process to fix recorded/filled cells from mouse and human brain slices with 4% PFA/2.5% Glutaraldehyde in PBS for future histochemical processing.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Post Patch Clamp Slice Fixation. protocols.io dx.doi.org/10.17504/protocols.io.bep3jdqn Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol provides step by step instructions for performing single-cell sorting by using BD FACSAria II machine or BD FACSAria Fusion machine and BD FACSDiva software.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. FACS Single Cell Sorting. protocols.io dx.doi.org/10.17504/protocols.io.bdvsi66e Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes how to isolate tissue from various ROI’s of the mouse brain followed by preparation of a highly concentrated single cell suspension for transcriptomic profiling on 10x Genomics platform.
Proper citation: Allen Institute for Brain Science 2020. Mouse Whole Cell Tissue Processing for 10x Genomics Platform. protocols.io dx.doi.org/10.17504/protocols.io.bc6bizan Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used to prepare 4% Paraformaldehyde (PFA) + 2.5% Glutaraldehyde Fixative. This is utilized to fix the tissue after electrophysiology recordings. Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. 4% PFA + 2.5% Glutaraldehyde Fixative. protocols.io dx.doi.org/10.17504/protocols.io.bg5sjy6e Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the procedures for intracardiac perfusion fixation of postnatal mice, including anesthesia, exsanguination, fixation, brain removal and post-fixation storage. Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Mouse Cardiac Perfusion Fixation and Brain Collection. protocols.io dx.doi.org/10.17504/protocols.io.bg5vjy66 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the procedure for transcardial perfusion of an adult mouse, followed by dissection, embedding, slicing, and blockface imaging of the mouse brain using either a Leica vibratome or a compresstome. The output tissue is suitable for use in electrophysiological recordings and/or live cell filling applications.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Slice Preparation and Blockface Imaging for Electrophysiology - Mouse. protocols.io dx.doi.org/10.17504/protocols.io.be2gjgbw Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocols provides detailed instructions for procedures needed for maintenance of the TissueCyte scanning systems.
Proper citation: Allen Institute for Brain Science 2020. Maintenance of TissueCyte System. protocols.io dx.doi.org/10.17504/protocols.io.bbx6ipre Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used to prepare 0.1M Phosphate Buffer. 0.1M Phosphate Buffer is used in the preparation of Sucrose solutions for cryoprotection.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. 0.1M Phosphate Buffer. protocols.io dx.doi.org/10.17504/protocols.io.bg43jyyn Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the delivery of a neuronal tracer using the Nanoject II. The surgery uses a stereotaxic system to target specific brain coordinates in the mouse. Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Injection of Viral Tracers by Nanoject. protocols.io dx.doi.org/10.17504/protocols.io.bd8qi9vw Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used to prepare Tris Buffered Saline (TNT). 10X TNT is the stock solution used to prepare 1X TNT, which equilibrates tissue and maintains pH in physiological range.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Tris Buffered Saline (TNT). protocols.io dx.doi.org/10.17504/protocols.io.bgncjvaw Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: Freshly prepared 4% Paraformaldehyde in PBS is used to fix adult and developing mouse brains during the transcardial perfusion process, as well as for immersion fixation. This solution can be used for no more than one week post-preparation date.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Fresh 4% Paraformaldehyde in PBS. protocols.io dx.doi.org/10.17504/protocols.io.bezijf4e Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the process for diaminobenzidine (DAB) detection of biocytin filled cells. This protocol is optimized for use with brain slices cut at 350 µm thick, in which cells are first filled with biocytin (i.e., post-electrophysiological recording), fixed in 4% PFA/2.5% glutaraldehyde, and transferred to PBS until ready to stain.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. DAB Detection of Biocytin Labeled Tissue. protocols.io dx.doi.org/10.17504/protocols.io.bep5jdq6 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol provides step by step instructions for performing single-cell sorting by using BD FACSAria II machine or BD FACSAria Fusion machine and BD FACSDiva software.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. FACS Single Cell Sorting. protocols.io dx.doi.org/10.17504/protocols.io.besbjean Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes general procedures used for subcutaneous injection in adult mice.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Subcutaneous Injection Adult Mouse. protocols.io dx.doi.org/10.17504/protocols.io.bezdjf26 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: Protocol is used for the manufacture of glass pipettes for patch clamp electrophysiology using the DMZ-Zeitz Universal puller.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Pulling Pipettes using the DMZ-Zeitz Universal. protocols.io dx.doi.org/10.17504/protocols.io.bdpfi5jn Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used for the degassing of critical reagents used in both the colorimetric and fluorescent in situ hybridization (ISH). This is crucial in eliminating hybridization bubbles from forming on the tissue. Hybridization bubbles result in no expression where they are located on the tissue.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. Degassing Reagents. protocols.io dx.doi.org/10.17504/protocols.io.bg4wjyxe Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used for the rapid generation of 3’ transcriptomic-NGS-ready- single-cell-libraries from pools of cells.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number's 1U01MH114812-01 and 1U19MH114830-01. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2021. 10Xv3.1 Genomics Sample Processing. protocols.io dx.doi.org/10.17504/protocols.io.brt5m6q6 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes bioanalyzer operation. The Agilent Bioanalyzer is used as a quality control step for PCR and IVT products. Peak size, shape, and sample concentration are recorded and used to evaluate samples on a pass/fail level.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Proper citation: Allen Institute for Brain Science 2020. 2100 Bioanalyzer Operation for PCR, GTP and IVT Products. protocols.io dx.doi.org/10.17504/protocols.io.bfzjjp4n Copy
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