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Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: Protocol allows for 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 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 2021. 10xV3 Genomics Sample Processing Protocol. protocols.io dx.doi.org/10.17504/protocols.io.bq7cmziw Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used to prepare 0.9% Saline Solution. 0.9% Sterile Saline solution is used as an initial rinse for the brains of mice prior to perfusion. 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. 0.9% Saline Solution. protocols.io dx.doi.org/10.17504/protocols.io.bg44jyyw 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.bdpji5kn Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used to produce AAV of any serotype with high purity and high titer.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. Production of AAV Virus by Iodixanol Gradient Ultracentrifugation. protocols.io dx.doi.org/10.17504/protocols.io.bdvxi67n Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used to produce crude preps of AAV of any serotype.
Proper citation: Allen Institute for Brain Science 2019. Production of Crude AAV Virus Extract. protocols.io dx.doi.org/10.17504/protocols.io.8djhs4n Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used to produce AAV of any serotype with high purity and high titer.
Proper citation: Allen Institute for Brain Science 2019. Production of AAV Virus by Iodixanol Gradient Ultracentrifugation. protocols.io dx.doi.org/10.17504/protocols.io.8dmhs46 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This SOP provides instruction to prepare Internal Solution which is used for patch clamp electrophysiology, modified for cleanliness and mRNA capture and sustainability.
Proper citation: Allen Institute for Brain Science 2019. Patch-Seq Internal Solution with Biocytin. protocols.io dx.doi.org/10.17504/protocols.io.9gih3ue Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used for in vitro transcription (IVT) reaction generating DIG-labeled anti-sense (AS) RNA from polymerase chain reaction (PCR) product for in situ hybridization (ISH), as well as fluorescent quantification of products using Ribogreen immediately following. This protocol is written for 96 reactions but can also be used for single reactions or in multiples of 96 by adjusting the amount of materials and equipment appropriately.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. Digoxigenin Labeled In Vitro Transcription and Ribogreen Quantification. protocols.io dx.doi.org/10.17504/protocols.io.bg4vjyw6 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the instructions for setup and scanning of fluorescently-labeled tissue sections mounted in agarose and attached to a slide using the TissueCyte 1000 block face 2-photon system.
Proper citation: Allen Institute for Brain Science 2020. Core Protocol for Serial Imaging of Fluorescently-Labeled Mouse Brain on the TissueCyte 1000 System. protocols.io dx.doi.org/10.17504/protocols.io.bbx4ipqw Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: 3, 3´ Diaminobenzidine (DAB) is a commonly used chromogen for histological staining. In the presence of peroxidase enzyme, DAB produces a brown precipitate that is insoluble in alcohol and xylene. Peroxidase catalyzes hydrogen peroxide oxidation of substrate DAB. This SOP provides instructions to prepare four different formulations of DAB. DAB for Immunohistochemical staining (IHC), DAB for Acetylcholinesterase (AChE) Staining, Nickel Enchanced DAB for Biocytin staining, and DAB for Biocytin staining.
Proper citation: Allen Institute for Brain Science 2020. DAB Solutions. protocols.io dx.doi.org/10.17504/protocols.io.bctkiwkw 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.beudjes6 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: Artificial Cerebrospinal Fluid III (ACSF.III) is used for applications including tissue bath solution during electrophysiological recording.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. Artificial Cerebrospinal Fluid III (ACSF.III). protocols.io dx.doi.org/10.17504/protocols.io.bdpci5iw Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the preparation of 1x3 glass slides for mounting embedded mouse brain samples for subsequent imaging and sectioning on TissueCyte 1000. 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. Slide Preparation for Mounting Samples for TissueCyte 1000 Sectioning and Imaging. protocols.io dx.doi.org/10.17504/protocols.io.bdvni65e Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used to prepare Phosphate Buffered Saline (PBS). PBS (-) and PBS (+) which includes Tween are used as biologically neutral wash solutions in many processes including In Situ Hybridization (ISH), Immunohistochemistry (IHC) and Fixation.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. Phosphate Buffered Saline (PBS). protocols.io dx.doi.org/10.17504/protocols.io.bezgjf3w Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes PCR1 and PCR2 reactions beginning with cDNA to generate transcript templates ready for IVT reaction. This protocol is written for 192 (2x96) reactions but can also be used for fewer or more reactions by adjusting the amount of materials and equipment appropriately.
Proper citation: Allen Institute for Brain Science 2020. PCR Prep from cDNA for IVT Reactions. protocols.io dx.doi.org/10.17504/protocols.io.bcsfiwbn Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes making Artificial Cerebrospinal Fluid IV (ACSF.IV). ACSF.IV is used for multiple applications including incubation of fresh mouse brain slices prior to electrophysiological recording.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. Artificial Cerebrospinal Fluid IV (ACSF.IV). protocols.io dx.doi.org/10.17504/protocols.io.bdpbi5in Copy
Authors: ZengU19 BRAIN grant
Group: BICCN, Allen Institute for Brain Science
Summary: Protocol to generate full-length cDNA from single cells, or nuclei, using Takara SMARTer V4.
Proper citation: ZengU19 BRAIN grant 2019. SMARTerV4 (1x) Amplification for single-cell or single-nuclei RNASeq Protocol. protocols.io dx.doi.org/10.17504/protocols.io.7d6hi9e Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol outlines strategies to solve common problems that occur when setting up the TissueCyte 1000 system for normal operation.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. TissueCyte Troubleshooting Guide. protocols.io dx.doi.org/10.17504/protocols.io.bdpsi5ne Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes PCR1 and PCR2 reactions beginning with cDNA to generate transcript templates ready for IVT reaction. This protocol is written for 192 (2x96) reactions but can also be used for fewer or more reactions by adjusting the amount of materials and equipment appropriately.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. PCR Prep from cDNA for IVT Reactions. protocols.io dx.doi.org/10.17504/protocols.io.bdpni5me Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the process to obtain electrophysiological recordings and cellular contents from neurons in postnatal mouse and/or human brain slices.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. Patch-Seq Recording and Extraction. protocols.io dx.doi.org/10.17504/protocols.io.bdpei5je Copy
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