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Authors: Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: This SOP describes the methods used to ship paraffin-embedded blocks to the University of Zurich.
Proper citation: Marda Jorgensen 2020. Shipping Paraffin Blocks to the Bodenmiller Lab for IMC Analysis. protocols.io dx.doi.org/10.17504/protocols.io.bf5njq5e Copy
Authors: Franchesca Farris, Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: This is an appendix for spleen SOP.
Proper citation: Franchesca Farris, Marda Jorgensen 2020. SOP Appendix for Spleen. protocols.io dx.doi.org/10.17504/protocols.io.bcwhixb6 Copy
Authors: Marda Jorgensen, Michelle Daniel
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Version 11 of the antibody labelling protocol published by Fluidigm.
Proper citation: Marda Jorgensen, Michelle Daniel 2020. Maxpar Antibody Labeling for Imaging Mass Cytometry. protocols.io dx.doi.org/10.17504/protocols.io.bf5jjq4n Copy
Authors: Marda Jorgensen, Seth Currlin
Group: Optical Clearing of Tissue, Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: The Zeiss Lightsheet 7 is used to quickly and efficiently scan large tissue volumes using laser lightsheet illumination. The focused laser sheets and imaging plane are static while the tissue is moved throughout the selected volume for three dimensional image acquisition. This protocol will cover the procedures used for the day of lightsheet image acquisition.Setting the acquisition volume: Large scanning volumes are composed of multiple z-stack being stitched back together based on overlapping edges of the scanning window. This requires setting an overall z-stack, which must include the entirety of tissue as covered by adjoining z-stacks. This becomes tricky when considering irregularly shaped tissues and their orientation relative to the various laser and imaging objectives.Dual-side lightsheet alignment: To obtain a strong and clear image from large tissues it is helpful to illuminate the sample with both the left and right laser. Each lasersheet can be angled slightly within the focal plane to account for differences in the R.I. of the tissue and R.I. media (TDE). This is a manual tuning process and is not intended for major R.I. differences.Protocol Overview:- Refractive index matching of cleared tissue samples. - Mounting tissue onto sample holder.- Image acquisition in Zen 3.1 LS (black edition), including: - Setting the acquisition volume - Dual-sided lightsheet alignment
Proper citation: Marda Jorgensen, Seth Currlin 2020. Lightsheet Microscope Acquisition Protocol. protocols.io dx.doi.org/10.17504/protocols.io.begajbse Copy
Authors: Jerelyn Nick, Marda Jorgensen, Seth Currlin
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 Light Sheet Fluorescence Microscopy 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: Jerelyn Nick, Marda Jorgensen, Seth Currlin 2020. HuBMAP TMC-Florida/Zurich Light Sheet Fluorescence Microscopy Modality Overview. protocols.io dx.doi.org/10.17504/protocols.io.bqaamsae Copy
Authors: Michelle Daniel, Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: This SOP describes the preparation of compensation slides with single antibodyconjugatespots that can be used to estimate mass channel spillover for IMC. Spillovervaries from metal lot to lot due to varying impurities. Once a spillover matrix for a specificset of antibody-conjugates has been generated, the experiment of interest, in which thesame antibody conjugates have been used, can be spillover-corrected.It is crucial that this procedure is performed for the exact same antibody conjugates thatare used to pipette the antibody panel for sample staining.The original publication can be viewed here:“Compensation of Signal Spillover in Suspension and Imaging Mass Cytometry” by S.Chevrier, HL Crowell and VRT Zanotelli et al., Cell Systems, 2018
Proper citation: Michelle Daniel, Marda Jorgensen 2020. Imaging Mass Cytometry Compensation Slide Preparation. protocols.io dx.doi.org/10.17504/protocols.io.bf2djqa6 Copy
Authors: Marda Jorgensen, 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 OCT embedding of HuBMAP fresh frozen specimens.
Proper citation: Marda Jorgensen, Jerelyn Nick 2020. HuBMAP: Embedding Fresh Frozen OCT Samples. protocols.io dx.doi.org/10.17504/protocols.io.bcwsixee Copy
Authors: Franchesca Farris, Marda Jorgensen, Leigh Propper
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: This protocol will detail our method of utilizing the H&E stain as a nonspecific entity to achieve optimal results in our staining process for our research samples.Hematoxylin and Eosin stains are used in many areas of the laboratory, including frozen sections, fine needle aspirates, and paraffin fixed embedded tissues. There are multiple factors involved in determining what makes a well stained slide, and one of the primary factors involved in the final product is pathologist preference. Other important factors also play important roles in your final stain, such as reagents and their quality, consistency, initial microtomy quality and selecting a sound procedure. 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 brought out 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: Franchesca Farris, Marda Jorgensen, Leigh Propper 2019. UF / HuBMAP - H&E Staining Process . protocols.io dx.doi.org/10.17504/protocols.io.9qbh5sn Copy
Authors: Marda Jorgensen, Jerelyn Nick
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: This is an appendix diagram for spleen tissue cutting.
Proper citation: Marda Jorgensen, Jerelyn Nick 2020. Appendix for Spleen SOP. protocols.io dx.doi.org/10.17504/protocols.io.bbcviiw6 Copy
Authors: Marda Jorgensen, Maigan Brusko
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Proper citation: Marda Jorgensen, Maigan Brusko 2020. HubMap UF TMC Tissue Dissociation to Single Cell. protocols.io dx.doi.org/10.17504/protocols.io.bd9vi966 Copy
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