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Authors: Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: The purpose of this Standard Operating Procedure is to outline procedures for processing and storing spleen received for HuBMAP consortium assay and analysis.
Proper citation: Marda Jorgensen 2020. Case Processing SOP (Spleen). protocols.io dx.doi.org/10.17504/protocols.io.bc3kiykw Copy
Authors: Elizabeth Neumann, Jamie Allen, David Anderson, Danielle Gutierrez, Jeff Spraggins
Group: VU Biomolecular Multimodal Imaging Center, Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Scope:Acquire 10 μm spatial resolution imaging mass spectrometry (IMS) datasets of ~ 5mm2 regions of tissue.Expected Outcome:A series of ion images that can be used to visualize the distribution of lipids localizing to physiological regions within the human kidney.
Proper citation: Elizabeth Neumann, Jamie Allen, David Anderson, Danielle Gutierrez, Jeff Spraggins 2019. High Resolution Imaging Mass Spectrometry Analysis using Bruker Daltonics Platforms. protocols.io dx.doi.org/10.17504/protocols.io.7gdhjs6 Copy
Authors: Jamie Allen, Jennifer Harvey, Maya Brewer, Mark De Caestecker, Jeff Spraggins
Group: VU Biomolecular Multimodal Imaging Center, Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Scope:The PAS stain is used to demonstrate polysaccharides such as glycogen, and mucosubstances such as glycoproteins, glycolipids and mucins in tissues. It is used as a replacement for the H&E in kidney pathology.Expected Outcome:Intermyofibrillar Network…………………………..Pink to RoseGlycogen………………………………………………….…..Pink to RoseMyofibrils……………………………………………………..UnstainedType I fibers………………………………………………….LighterType II fibers…………………………………………………DarkerBlood Vessel walls & connective tissue…..Faintly Stained
Proper citation: Jamie Allen, Jennifer Harvey, Maya Brewer, Mark De Caestecker, Jeff Spraggins 2019. PAS Staining of Fresh Frozen or Paraffin Embedded Human Kidney Tissue. protocols.io dx.doi.org/10.17504/protocols.io.4qngvve Copy
Authors: Marda Jorgensen, Jerelyn Nick, Franchesca Farris, Jesus Penaloza
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: This protocol is an overview of all of the protocols currently in use for the CODEX modality at HubMAP Tissue Mapping Center,TMC-Florida/Zurich. It includes protocols.io links to each of the individual protocols that make up this project workflow, from processing tissue and imaging the samples, to uploading the final data to the HuBMAP HIVE.
Proper citation: Marda Jorgensen, Jerelyn Nick, Franchesca Farris, Jesus Penaloza 2020. HuBMAP TMC-Florida/Zurich CODEX Modality Overview. protocols.io dx.doi.org/10.17504/protocols.io.be9pjh5n Copy
Authors: Elizabeth Neumann, Jamie Allen, Heath Patterson, Danielle Gutierrez, Jeff Spraggins
Group: VU Biomolecular Multimodal Imaging Center, Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Scope: Obtain autofluorescence microscopy images of tissues after IMS analysisExpected Outcome: A brightfield and green autofluorescence microscopy image of the tissue section that enables registration and correlation of different imaging modalities on a pixel by pixel basis.
Proper citation: Elizabeth Neumann, Jamie Allen, Heath Patterson, Danielle Gutierrez, Jeff Spraggins 2020. Post-IMS Autofluorescence Microscopy. protocols.io dx.doi.org/10.17504/protocols.io.879hzr6 Copy
Authors: Seth Currlin, Marda Jorgensen
Group: Optical Clearing of Tissue, Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: In order to track tissue position and orientation each sample is photographed when received for lightsheet microscopy (Figure 1, A). Large samples (>1 cm3), such as the spleen in Figure 1, are cut into 2 mm sections (Figure 1, B) to facilitate tissue clearing, staining, and three-dimensional imaging. These tissues are batch processed and withdrawn from the lipid clearing pipeline as needed (Figure 1, C); at this time a novel identifier is assigned to the sample.
Proper citation: Seth Currlin, Marda Jorgensen 2020. Lightsheet Tissue Intake - Photodocumentation and Tracking. protocols.io dx.doi.org/10.17504/protocols.io.bef6jbre Copy
Authors: Marda Jorgensen, Franchesca Farris
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Hematoxylin and Eosin stains are the prefered method for histopathologic assessment of tissue sections.Hematoxylin is used to illustrate nuclear detail in cells. The resulting intensity and shade of the dye in the cells is primarily reliant on the length of time the sample spends in hematoxylin. The cytoplasmic component of the section is identified by Eosin, which is the most commonly used counterstain in histology. It's varying shades of pink and red provide an exceptional contrast between the nuclear and cytoplasmic areas of interest.
Proper citation: Marda Jorgensen, Franchesca Farris 2020. UF H&E Staining. protocols.io dx.doi.org/10.17504/protocols.io.beamjac6 Copy
Authors: Shin Lin, Yiing Lin
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Proper citation: Shin Lin, Yiing Lin 2020. Protocol for Freezing in OCT. protocols.io dx.doi.org/10.17504/protocols.io.bpx7mprn Copy
Authors: Jerelyn Nick, Franchesca Farris, Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Standard IHC staining, such as this DAB protocol, is included in the validation of new antibodies at the University of Florida Tissue Mapping Center, TMC-Florida/Zurich. This procedure includes all of the reagents necessary to perform a DAB based immunohistochemistry reaction. DAB Substrate (3,3’-diaminobenzidine) produces a brown reaction product in the presence of peroxidase (HRP) enzyme.
Proper citation: Jerelyn Nick, Franchesca Farris, Marda Jorgensen 2020. DAB Staining of FFPE Slides. protocols.io dx.doi.org/10.17504/protocols.io.bdiei4be Copy
Authors: Elizabeth Neumann, Heath Patterson, Jamie Allen, Maya Brewer, Mark De Caestecker, Danielle Gutierrez, Jeff Spraggins
Group: VU Biomolecular Multimodal Imaging Center, Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Scope:Obtain autofluorescence microscopy images of tissues.Expected Outcome:An RGB autofluorescence microscopy image of the tissue section that enables registration and correlation of different imaging modalities on a pixel by pixel basis.
Proper citation: Elizabeth Neumann, Heath Patterson, Jamie Allen, Maya Brewer, Mark De Caestecker, Danielle Gutierrez, Jeff Spraggins 2019. Autofluorescence Microscopy Data Acquisition. protocols.io dx.doi.org/10.17504/protocols.io.7e3hjgn Copy
Authors: Leigh Propper, Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: This method details our microtomy (sectioning) process for reasearch specimens involved with HuBMAP.This process follows the microtomy process that follows after receiving, processing, and embedding of research specimens into formalin fixed, paraffin embedded tissue blocks.
Proper citation: Leigh Propper, Marda Jorgensen 2019. HuBMAP Tissue Sectioning for FFPE Specimens. protocols.io dx.doi.org/10.17504/protocols.io.bahdib26 Copy
Authors: Marda Jorgensen, Franchesca Farris, Jerelyn Nick
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: The purpose of this Standard Operating Procedure (SOP) is to outline procedures for the automated processing of formalin fixed HuBMAP specimens for paraffin embedding.
Proper citation: Marda Jorgensen, Franchesca Farris, Jerelyn Nick 2020. HuBMAP - Tissue Processing with Paraffin Infiltration. protocols.io dx.doi.org/10.17504/protocols.io.8nzhvf6 Copy
Authors: Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: The purpose of this Standard Operating Procedure is to outline procedures for processing and storing spleen received for HuBMAP consortium assay and analysis.
Proper citation: Marda Jorgensen 2020. Case Processing SOP (Spleen). protocols.io dx.doi.org/10.17504/protocols.io.bbcuiiww Copy
Authors: Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Method for sectioning paraffin blocks for staining
Proper citation: Marda Jorgensen 2019. FFPE Tissue Sectioning for Staining. protocols.io dx.doi.org/10.17504/protocols.io.8kvhuw6 Copy
Authors: Anup Sood, Eric Williams, Liz McDonough
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community, GE Research
Summary: ScopeThe purpose of this protocol is to conjugate purified antibodies to Cy dyes. Antibody conjugation involves combining dyes with antibodies to achieve a fluorescently labeled antibody. Each antibody will be labeled with a molar equivalent of dye; the quantitative output of this is deemed D/P. In the initial conjugation step, each antibody will be labeled with differing D/P levels and validated to determine which level is optimal.
Proper citation: Anup Sood, Eric Williams, Liz McDonough 2021. Cell DIVE™ Platform | Ab Conjugation: Initial Conjugation & Scale up Conjugation. protocols.io dx.doi.org/10.17504/protocols.io.bp55mq86 Copy
Authors: Jamie Allen, Elizabeth Neumann, Jeff Spraggins
Group: VU Biomolecular Multimodal Imaging Center, Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Scope:To describe the procedure for spraying tissue sections with DAN for imaging lipids.Expected Outcome:Slides should be coated with DAN. Tissue sections should be imaged within 24 hours of matrix application.
Proper citation: Jamie Allen, Elizabeth Neumann, Jeff Spraggins 2019. Automatic Deposition of DAN Matrix using a TM Sprayer for MALDI Analysis of Lipids. protocols.io dx.doi.org/10.17504/protocols.io.4srgwd6 Copy
Authors: Michelle Daniel, Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: The following SOP describes procedures required for the antibody staining of formalin fixed,paraffin-embedded (FFPE) samples. In this SOP, the term “samples” refers to FFPEtissue samples.
Proper citation: Michelle Daniel, Marda Jorgensen 2020. Imaging Mass Cytometry Antibody Staining . protocols.io dx.doi.org/10.17504/protocols.io.bfz8jp9w Copy
Authors: Leigh Propper, Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: This method details our microtomy (sectioning) process for reasearch specimens involved with HuBMAP.This process follows the microtomy process that follows after receiving, processing, and embedding of research specimens into formalin fixed, paraffin embedded tissue blocks.
Proper citation: Leigh Propper, Marda Jorgensen 2019. HuBMAP Tissue Sectioning for FFPE Specimens. protocols.io dx.doi.org/10.17504/protocols.io.93qh8mw Copy
Authors: Jamie Allen, Jeff Spraggins, Danielle Gutierrez
Group: VU Biomolecular Multimodal Imaging Center, Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Scope:To describe the procedure for the lysis, reduction/alkylation, trypsin digestion, and clean-up of tissue. Lysis will cover the lysing of tissue and protein concentration. Acetone precipitation will cover the precipitation of proteins. Digestion will cover the process for digesting 100 µg of protein using the Agilent AssayMap Bravo Robot and Promega Rapid Trypsin/LysC. The clean-up of the cells will cover the desalting process on the Agilent AssayMap Bravo and the subsequent preparation of the samples for LC/MS peptide analysis.Expected Outcome/Data:Cell samples lysed, digested, and desalted for analysis on MS instrument. Samples to be analyzed within one or two days of desalting.
Proper citation: Jamie Allen, Jeff Spraggins, Danielle Gutierrez 2019. Automated, Rapid Preparation of Tissue Sections for Proteomic Analysis. protocols.io dx.doi.org/10.17504/protocols.io.67nhhme Copy
Authors: Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: This document outlines the required criteria for inclusion for donors of thymus, lymph nodes and spleen in the HuBMAP project.
Proper citation: Marda Jorgensen 2020. Donor Acceptance Criteria for TMC Florida/Zurich HuBMAP Inclusion. protocols.io dx.doi.org/10.17504/protocols.io.bipykdpw Copy
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