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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.
Proper citation: Allen Institute for Brain Science 2020. Post Patch Clamp Slice Fixation. protocols.io dx.doi.org/10.17504/protocols.io.bctciwiw Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the procedure for receiving human surgical tissue, followed by trimming, mounting, embedding, and slicing under sterile conditions in order to produce tissue suitable for both slice culture as well as acute slice electrophysiology recording.
Proper citation: Allen Institute for Brain Science 2020. Human Neurosurgical Tissue Processing. protocols.io dx.doi.org/10.17504/protocols.io.bctdiwi6 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the preparation of a fixed mouse brain specimen bonded to an agarose block for blockface imaging and sectioning using the TissueCyte 1000 system.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 Specimen Embedding Acrylamide Coembedding. protocols.io dx.doi.org/10.17504/protocols.io.ber4jd8w Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: 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. 10Xv2 RNASeq Sample Processing. protocols.io dx.doi.org/10.17504/protocols.io.bq68mzhw Copy
Authors: Brian Lee, Kristen Hadley, Allen Institute for Brain Science
Group: Allen Institute for Brain Science
Summary: This protocol is a detailed version of the Allen Institute's Patch-Seq Recording and Extraction protocol, which 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: Brian Lee, Kristen Hadley, Allen Institute for Brain Science 2020. Patch-Seq Recording and Extraction Detailed Protocol. protocols.io dx.doi.org/10.17504/protocols.io.bpbuminw Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes how to make Artificial Cerebrospinal Fluid VIII (ACSF.VIII). ACSF.VIII is used for multiple applications including incubation of fresh human 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 VIII (ACSF.VIII). protocols.io dx.doi.org/10.17504/protocols.io.bdpai5ie Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: The protocol describes how 1x3 glass microscope slides are coated with gelatin to improve adherence of fixed adult mouse brain sections or other tissue to slides.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. Coating Slides with Gelatin. protocols.io dx.doi.org/10.17504/protocols.io.bf64jrgw 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.bepzjdp6 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.
Proper citation: Allen Institute for Brain Science 2019. FACS Single Cell Sorting. protocols.io dx.doi.org/10.17504/protocols.io.bambic2n 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.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. Maintenance of TissueCyte System. protocols.io dx.doi.org/10.17504/protocols.io.ber3jd8n Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: Isolation of nuclei from frozen adult human brain tissue or thawed and microdissected brain tissue sections for FPCR and/or RNA-seq analysis. 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 2021. Isolation of Nuclei from Adult_Human_Brain_Tissue. protocols.io dx.doi.org/10.17504/protocols.io.bq7dmzi6 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol is used to prepare 20% Paraformaldehyde (PFA) in 5X Phosphate-buffered saline (PBS). 20% PFA in 5X PBS is utilized to make the 4% PFA in 1X PBS reagent.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. 20% Paraformaldehyde in 5X PBS. protocols.io dx.doi.org/10.17504/protocols.io.bg4zjyx6 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.be2djga6 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: Isolation of nuclei from frozen adult human brain tissue or thawed and microdissected brain tissue sections for RNA-seq analysis. 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 2021. Isolation of Nuclei from Adult Human Brain Tissue for 10x Genomics Platform. protocols.io dx.doi.org/10.17504/protocols.io.bq64mzgw Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the synthesis of first-strand cDNA from total RNA using the Invitrogen SuperScript III Reverse Transcriptase system.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. cDNA Synthesis. protocols.io dx.doi.org/10.17504/protocols.io.bgnbjvan Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: 50 mM Phosphate Buffer can be used as a general buffer solution for multiple applications. It is suitable for use as a buffer for specimen embedding, as part of brain preparation associated with TissueCyte 2P Serial Imaging, described in PF0315 TissueCyte Specimen Embedding Acrylamide Coembedding.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. 50 mM Phosphate Buffer (with 0.02% sodium azide option). protocols.io dx.doi.org/10.17504/protocols.io.bezjjf4n Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the synthesis of first-strand cDNA from total RNA using the Invitrogen SuperScript III Reverse Transcriptase system.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. cDNA Synthesis. protocols.io dx.doi.org/10.17504/protocols.io.besejebe Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol provides guidance in the assessment of quality of tissues post perfusion and dissection and applies to all such procedures. Perfusion and dissection quality will be categorized on the following scale: perfect, good, fair, and poor. This SOP also presents written descriptions, and images, of specific category attributes. 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. Tissue Quality Evaluation for Brain Perfusion/Dissection Specimens. protocols.io dx.doi.org/10.17504/protocols.io.bd8yi9xw 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.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 Internal Solution with Biocytin. protocols.io dx.doi.org/10.17504/protocols.io.besdjea6 Copy
Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the system that is used to capture Z stack images which are reassembled post-processing into 3D reconstructions.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. Bright Field Image Capture with Zeiss AxioImager. protocols.io dx.doi.org/10.17504/protocols.io.bdpmi5k6 Copy
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