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Authors: Nabil-Fareed Alikhan, Emma Griffiths, Ruth Timme, Duncan MacCannell
Group: Coronavirus Method Development Community, PHA4GE
Summary: Guidance on how to populate the extended PHA4GE metadata package for SARS-CoV-2 submissions, maximizing interoperability for covid-19 surveillance.
Proper citation: Nabil-Fareed Alikhan, Emma Griffiths, Ruth Timme, Duncan MacCannell 2020. SOP for populating EBI submission templates (ENA). protocols.io dx.doi.org/10.17504/protocols.io.bh5dj826 Copy
Authors: Prakash S Bisen
Summary: Humus is a complex mixture of heterogeneous organic compounds derived from plant and animal residues and is formed by the reaction of decomposition, synthesis, and polymerization. It is an integral part of soil organic matter and can be extracted by neutral salts or alkali separating humic and nonhumic fraction. .
Proper citation: Prakash S Bisen 2015. Estimation of Humus in Soil/Manure. protocols.io dx.doi.org/10.17504/protocols.io.de63hd Copy
Authors: Shahzad S. Khan, Herschel S. Dhekne, Francesca Tonelli, Suzanne R. Pfeffer
Summary: We describe here our method for immunostaining of primary cilia in brain sections from wild type and LRRK2 or other mutant mice. Included are procedures for perfusion, tissue harvesting, sectioning and staining, image acquisition and analysis.
Proper citation: Shahzad S. Khan, Herschel S. Dhekne, Francesca Tonelli, Suzanne R. Pfeffer 2020. Analysis of Primary Cilia in Rodent Brain By Immunofluorescence Microscopy. protocols.io dx.doi.org/10.17504/protocols.io.bnwimfce Copy
Authors: Jeff Strohm, Robert Hanner, Richard J Heck
Summary: This protocol provides the sampling and molecular biology lab methods used to prepare microbial amplicons for MiSeq sequencing.
Proper citation: Jeff Strohm, Robert Hanner, Richard J Heck 2018. The influence of agricultural tillage practices on soil biodiversity: Soil metagenomic methods, microbial community. protocols.io dx.doi.org/10.17504/protocols.io.efjbbkn Copy
Authors: Jia Ren Lin, Benjamin Izar, Zoltan Maliga, Yu-An Chen, Giorgio Gaglia, Ziming Du, Clarence Yapp, Shaolin Mei, Sandro Santagata, Peter Sorger
Group: Laboratory of Systems Pharmacology, NCIHTAN
Summary: The architecture of normal and diseased tissues strongly influences the development and progression of disease as well as responsiveness and resistance to therapy. We describe a tissue-based cyclic immunofluorescence (t-CyCIF) method for highly multiplexed immunofluorescence imaging of specimens mounted on glass slides. t-CyCIF generates up to 60-plex images using an iterative process (a cycle) in which conventional low-plex fluorescence images are repeatedly collected from the same sample and then assembled into a high dimensional representation. t-CyCIF requires no specialized instruments or reagents and is compatible with super-resolution imaging; we demonstrate its application to quantifying signal transduction cascades, tumor antigens and immune markers in diverse tissues and tumors. The simplicity and adaptability of t-CyCIF makes it an effective method for pre-clinical and clinical research and a natural complement to single-cell genomics.Key resources, reagents, and software are listed at the HMS LINCS Center Publication Page http://lincs.hms.harvard.edu/lin-elife-2018/ (RRID:SCR_016370). This page provides links to an OMERO image database from which individual images can be obtained; stitched and registered image panels can be obtained at www.cycif.org (RRID:SCR_016267) and a video illustrating the t-CyCIF method can be found at https://vimeo.com/269885646.This protocol is used in the following manuscripts: Lin J-R, Izar B, Wang S, Yapp C, Mei S, Shah P, Santagata S, Sorger PK. (2018). Highly multiplexed immunofluorescence imaging of human tissues and tumors using t-CyCIF and conventional optical microscopes. eLife. PMID: 29993362 Du Z, Lin JR, Rashid R, Maliga Z, Wang S, Aster J, Izar B, Sorger PK, Santagata S. (2019). Qualifying antibodies for image-based immune profiling and multiplexed tissue imaging. Nature Protocols. PMID: 31534232The original t-CyCIF protocol can be found at dx.doi.org/10.17504/protocols.io.rpxd5pn.
Proper citation: Jia Ren Lin, Benjamin Izar, Zoltan Maliga, Yu-An Chen, Giorgio Gaglia, Ziming Du, Clarence Yapp, Shaolin Mei, Sandro Santagata, Peter Sorger 2020. Tissue Cyclic Immunofluorescence (t-CyCIF). protocols.io dx.doi.org/10.17504/protocols.io.bjiukkew Copy
Authors: BioLegend, Inc.
Group: BioLegend
Proper citation: BioLegend, Inc. 2016. Treg Polarization of Mouse CD4+ Cells. protocols.io dx.doi.org/10.17504/protocols.io.ex7bfrn Copy
Authors: Björn Brembs
Group: Brembs Lab
Summary: We use this protocol to test if flies have preferance for darkness or light (phototactic or non-phototactic response).
Proper citation: Björn Brembs 2015. T-maze phototaxis with LEDs. protocols.io dx.doi.org/10.17504/protocols.io.c76zrd Copy
Authors: Ruilin Tian, Jason Hong, Martin Kampmann
Group: Neurodegeneration Method Development Community, KampmannLab
Proper citation: Ruilin Tian, Jason Hong, Martin Kampmann 2019. iPSC editing with TALENs. protocols.io dx.doi.org/10.17504/protocols.io.8dahs2e Copy
Authors: Noah Toppings
Group: XPRIZE Rapid Covid Testing
Proper citation: Noah Toppings 2020. RT-LAMP Reaction. protocols.io dx.doi.org/10.17504/protocols.io.bkp7kvrn Copy
Authors: Adrieli Sachett, Matheus Gallas-Lopes, Greicy M M Conterato, Radharani , Ana Herrmann, Angelo Piato
Group: Fish behavior and physiology
Summary: Zebrafish are incresingly used as a model animal in neuroscience research. Here we describe a protocol to quantify nonprotein sulfhydryl groups (NPSH), an indirect evaluation of the levels of reduced glutathione (GSH), a major oxidative stress defense in the central nervous system.
Proper citation: Adrieli Sachett, Matheus Gallas-Lopes, Greicy M M Conterato, Radharani , Ana Herrmann, Angelo Piato 2020. Quantification of nonprotein sulfhydryl groups (NPSH) optimized for zebrafish brain tissue. protocols.io dx.doi.org/10.17504/protocols.io.bjrkkm4w Copy
Authors: LI-COR Biosciences
Group: LI-COR Biosciences
Summary: The Odyssey Fc Imager, with 600 channel capabilities, can image agarose gels stained with popular DNA stains, such as ethidium bromide and SYBR Safe DNA stain, with sub-nanogram sensitivity. The Odyssey Fc Imager contains a 532 nm diffuse source with an excitation maximum of 520 nm and a detection maximum of 600 nm. These instrument parameters are within the range of the excitation and emission wavelengths of ethidium bromide (Ex/Em = 302 & 518/605 nm) and other visible fluorescent nucleic acid stains and provide a sensitive gel documentation option.
SYBR Safe DNA stain (Ex/Em = 502/530 nm) has also been tested on the Odyssey Fc Imager (using the 600 channel) with sensitivities exceeding ethidium bromide detection. The maximum fluorescence emission wavelength of SYBR Safe is very close to the maximum excitation wavelength. However, the Odyssey Fc 600 channel collects excitation light at a wavelength 50 nm higher than the maximum excitation wavelength of SYBR Safe. These instrument properties decrease the background and improve the signal-to-noise ratio for nucleic acid detection.
Specific instructions are given in this protocols for ethidium bromide and SYBR Safe use. Other nucleic acid binding stains may also be compatible with the Odyssey Fc Imager. Please check the excitation and emission spectra of each stain.
Developed for: Odyssey Fc Imaging System
Proper citation: LI-COR Biosciences 2018. Image Acquisition on the Odyssey Fc Imager. protocols.io dx.doi.org/10.17504/protocols.io.guubwww Copy
Authors: Yumin Yang, Hongshen Wan
Proper citation: Yumin Yang, Hongshen Wan 2020. Protocol for early vigour QTL mapping. protocols.io dx.doi.org/10.17504/protocols.io.bgrnjv5e Copy
Authors: Paul Rutten, Richard Tennant, Jacob Beal, Christopher Workman, Traci Haddock-Angelli, Natalie Farny, Vinoo Selvarajah
Group: iGEM Measurement
Summary: This procedure can be used to calibrate OD600 to colony forming unit (CFU) counts, which are directly relatable to the cell concentration of the culture, i.e. viable cell counts per mL. This protocol assumes that 1 bacterial cell will give rise to 1 colony.
For the CFU protocol, you will need to count colonies for your two Positive Control (BBa_I20270) cultures and your two Negative Control (BBa_R0040) cultures. Protocol based on this Yeast Plate Count Protocol.
Proper citation: Paul Rutten, Richard Tennant, Jacob Beal, Christopher Workman, Traci Haddock-Angelli, Natalie Farny, Vinoo Selvarajah 2019. Calibration Protocol - Conversion of OD600 to Colony Forming Units (CFUs). protocols.io dx.doi.org/10.17504/protocols.io.5gjg3un Copy
Authors: Joao Vitor Molino
Summary: This protocols describe the steps required for obtain a growth curve of Chlamydomonas reinhardtii and fluorescent protein expression of mVenus and mCherry.
Proper citation: Joao Vitor Molino 2020. Growth curve for Chlamydomonas reinhardtii. protocols.io dx.doi.org/10.17504/protocols.io.bpvbmn2n Copy
Authors: Addgene The Nonprofit Plasmid Repository
Summary: This protocol is for DNA quantification. To see the full abstract and additional resources, please visit https://www.addgene.org/protocols/dna-quantification/.
Proper citation: Addgene The Nonprofit Plasmid Repository 2020. DNA Quantification. protocols.io dx.doi.org/10.17504/protocols.io.4g8gtzw Copy
Authors: New England Biolabs
Group: New England Biolabs (NEB)
Summary: This is the quick version of the Monarch® DNA Gel Extraction Kit Protocol (NEB #T1020). For the full protocol, please click here.
Proper citation: New England Biolabs 2016. Quick Protocol for Monarch® DNA Gel Extraction Kit (NEB #T1020). protocols.io dx.doi.org/10.17504/protocols.io.ejwbcpe Copy
Authors: Jing Xu
Proper citation: Jing Xu 2017. Fermentation and Extraction. protocols.io dx.doi.org/10.17504/protocols.io.jipckdn Copy
Authors: Daisuke Katagiri, Takamune Takahashi
Group: Diabetic Complications Consortium
Summary: Summary This protocol describes a protocol to evaluate histological fibrosis in mouse kidney. Diabetic Complication:References 1. Katagiri D, Hamasaki Y, Doi K, et al. Interstitial renal fibrosis due to multiple cisplatin treatments is ameliorated by semicarbazide-sensitive amine oxidase inhibition. Kidney Int 2015; 89: 374 - 385. 2. Hara S, Umeyama K, Yokoo T, et al. Diffuse glomerular nodular lesions in diabetic pigs carrying a dominant-negative mutant hepatocyte nuclear factor 1-alpha, an inheritant diabetic gene in humans. PLoS One 2014; 9: e92219.
Proper citation: Daisuke Katagiri, Takamune Takahashi 2019. Histological Evaluation of Renal Fibrosis in Mice. protocols.io dx.doi.org/10.17504/protocols.io.3gygjxw Copy
Authors: Angel Justiz-Vaillant, Belkis Ferrer-Cosme
Group: University of the West Indies, [email protected]
Proper citation: Angel Justiz-Vaillant, Belkis Ferrer-Cosme 2020. ELISA for measurement of serum macrophage migration inhibitory factor (MIF). . protocols.io dx.doi.org/10.17504/protocols.io.bj7ekrje Copy
Authors: E. Dale Abel
Group: Diabetic Complications Consortium
Summary: Summary:This protocol describes the procedure used by the DiaComp for cardiac performance, oxygen consumption and substrate metabolic rates in isolated working mouse hearts.Diabetic Complication:References:1. Belke, D.D., Larsen, T.S., Gibbs, E.M., Severson, D.L. 2001.Glucose metabolism in perfused mouse hearts overexpressing human GLUT-4 glucose transporter. Am J Physiol Endocrinol Metab.280:E420-7.2. Belke, D.D., Larsen, T.S., Gibbs, E.M., Severson, D.L. 2000.Altered metabolism causes cardiac dysfunction in perfused hearts from diabetic (db/db) mice. Am J Physiol Endocrinol Metab.279:E1104-13.3. Belke, D.D., Larsen, T.S., Lopaschuk, G.D., Severson, D.L. 1999.Glucose and fatty acid metabolism in the isolated working mouse heart. Am J Physiol.277:R1210-7.4. Larsen, T.S., Belke, D.D., Sas, R., Giles, W.R., Severson, D.L., Lopaschuk, G.D., Tyberg, J.V. 1999.The isolated working mouse heart: methodological considerations. Pflugers Arch.437:979-85.5. Belke D.D., Betuing S, Tuttle M.J., Graveleau C, Young M.E., Pham M., Zhang D., Cooksey R.C., McClain D.A., Litwin S.E., Taegtmeyer H., Severson D., Kahn C.R., Abel E.D. 2002. Insulin signaling coordinately regulates cardiac size, metabolism, and contractile protein isoform expression. J Clin Invest 109:629-639.6. Mazumder P.K., O’Neill B.T., Roberts M.W., Buchanan J., Yun U.J., Cooksey R.C., Boudina S., Abel E.D. 2004. Impaired Cardiac Efficiency and Increased Fatty Acid Oxidation in Insulin Resistant ob/ob mouse hearts. Diabetes. 53: 2366-2374.7. Suga H: Ventricular energetics. Physiol Rev 70:247-277, 1990
Proper citation: E. Dale Abel 2019. Phenotypic Characterization of the Working Heart. protocols.io dx.doi.org/10.17504/protocols.io.8a2hsge Copy
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