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Name Authors DOI Group Summary Associated Publications RRIDs used Affiliations External URL Version Publication Date Proper Citation Record Last Update
Nutrient deplete/replete algal culture for elemental analysis
 
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Amy Zimmerman 10.17504/protocols.io.fcdbis6 VERVE Net, Worden Lab Purpose: To evaluate the influence of nutrient availability on cellular elemental composition (quotas and stoichiometry) in marine picoeukaryotes.Elemental quotas and ratios are assessed under nutrient replete and deplete conditions at the same time to minimize potential variation between experiments. Cells from an exponentially growing culture are concentrated by centrifugation, washed and re-suspended in a small volume of nutrient deplete media. These concentrated cells are used to inoculate triplicate replete and deplete culture flasks at a starting density corresponding to early-exponential growth. Culture growth is monitored daily. When the mean growth rate of the nutrient deplete treatment is half or less of the replete treatment (GRDEP/GRREP Monterey Bay Aquarium Research Institute 1 2016 Amy Zimmerman 2016. Nutrient deplete/replete algal culture for elemental analysis. protocols.io dx.doi.org/10.17504/protocols.io.fcdbis6 2021-03-29 03:08:41
nf-vcf-cataloguer
 
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Israel Aguilar Ordoñez 10.17504/protocols.io.bkmzku76 Whole genome variation in 27 Mexican indigenous populations, demographic and biomedical insights 'nf-vcf-cataloguer' is a tool, implemented in Nextflow, that generates a general table description in TSV format of the description of each category and subgroup of a VCF with the extended annotation made by VEP. Furthermore, it plots each subset of the consequences of variants. Instituto Nacional de Medicina Genómica (INMEGEN) https://github.com/Iaguilaror/nf-VCF-cataloguer 1 2020 Israel Aguilar Ordoñez 2020. nf-vcf-cataloguer. protocols.io dx.doi.org/10.17504/protocols.io.bkmzku76 2021-03-29 03:08:41
CUT&Tag Data Processing and Analysis Tutorial
 
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Ye Zheng, Kami Ahmad, Steven Henikoff 10.17504/protocols.io.bjk2kkye This tutorial is designed for processing and analyzing CUT&Tag data following the Benchtop CUT&Tag V.3 protocol. The illustration data used in this tutorial is the profiling of histone modifications in the human lymphoma K562 cell line, but the tutorial is generally applicable to any chromatin protein, including transcription factors, RNA polymerase II, and epitope-tagged proteins. For reproducible analysis, this tutorial is also available on GitHub at https://yezhengstat.github.io/CUTTag_tutorial/. Henikoff S, Henikoff JG, Kaya-Okur HS, Ahmad K, Efficient chromatin accessibility mapping in situ by nucleosome-tethered tagmentation. eLife doi: 10.7554/eLife.63274 Fred Hutchinson Cancer Research Center, Fred Hutchinson Cancer Research Center, Fred Hutchinson Cancer Research Center https://yezhengstat.github.io/CUTTag_tutorial/ 1 2020 Ye Zheng, Kami Ahmad, Steven Henikoff 2020. CUT&Tag Data Processing and Analysis Tutorial. protocols.io dx.doi.org/10.17504/protocols.io.bjk2kkye 2021-03-29 03:08:41
EPPS recipe for dissolved cobalt analyses
 
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Randie Bundy 10.17504/protocols.io.evcbe2w This is a protocol for making a trace metal clean 0.5 mol L-1 EPPS (N-(2-hydroxyethyl)piperazine-N-(3-propanesulfonic acid) solution for dissolved cobalt analyses. Woods Hole Oceanographic Institution 1 2016 Randie Bundy 2016. EPPS recipe for dissolved cobalt analyses. protocols.io dx.doi.org/10.17504/protocols.io.evcbe2w 2021-03-29 03:08:40
Flow cytometry
 
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Yuan Yao, morigen 10.17504/protocols.io.p9ndr5e Yao Y, Ma Y, Chen X, Bade R, Lv C, Zhu R (2018) Absence of RstA results in delayed initiation of DNA replication in Escherichia coli. PLoS ONE 13(7): e0200688. doi: 10.1371/journal.pone.0200688 Inner Mongolia People's Hospital, Inner Mongolia University https://doi.org/10.1371/journal.pone.0200688 1 2018 Yuan Yao, morigen 2018. Flow cytometry. protocols.io dx.doi.org/10.17504/protocols.io.p9ndr5e 2021-03-29 03:08:40
Diplonema Genomic DNA isolation
 
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Binnypreet Kaur1, 2, Drahomíra Faktorová1, 2, , Priscila Peña-Diaz1 and Julius Lukeš1, 2 10.17504/protocols.io.hfyb3pw Julius Lukes E-mail: [email protected] 1 Institute of Parasitology, Biology Centre, Czech Academy of Sciences, 37005, České Budějovice, Czech Republic 2 Faculty of Sciences, University of South Bohemia, 37005, České Budějovice, Czech Republic, 1 Institute of Parasitology, Biology Centre, Czech Academy of Sciences, 37005, České Budějovice, Czech Republic 2 Faculty of Sciences, University of South Bohemia, 37005, České Budějovice, Czech Republic, 1 Institute of Parasitology, Biology Centre, Czech Academy of Sciences, 37005, České Budějovice, Czech Republic 2 Faculty of Sciences, University of South Bohemia, 37005, České Budějovice, Czech Republic, 1 Institute of Parasitology, Biology Centre, Czech Academy of Sciences, 37005, České Budějovice, Czech Republic 2 Faculty of Sciences, University of South Bohemia, 37005, České Budějovice, Czech Republic, 1 Institute of Parasitology, Biology Centre, Czech Academy of Sciences, 37005, České Budějovice, Czech Republic 2 Faculty of Sciences, University of South Bohemia, 37005, České Budějovice, Czech Republic, 1 Institute of Parasitology, Biology Centre, Czech Academy of Sciences, 37005, České Budějovice, Czech Republic 2 Faculty of Sciences, University of South Bohemia, 37005, České Budějovice, Czech Republic, 1 Institute of Parasitology, Biology Centre, Czech Academy of Sciences, 37005, České Budějovice, Czech Republic 2 Faculty of Sciences, University of South Bohemia, 37005, České Budějovice, Czech Republic 1 2018 Binnypreet Kaur1, 2, Drahomíra Faktorová1, 2, , Priscila Peña-Diaz1 and Julius Lukeš1, 2 2018. Diplonema Genomic DNA isolation. protocols.io dx.doi.org/10.17504/protocols.io.hfyb3pw 2021-03-29 03:08:40
Glucose "Clamp" for measurement of whole body insulin-stimulated glucose disposal
 
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Donald McClain 10.17504/protocols.io.7s9hnh6 Diabetic Complications Consortium This is the standard protocol for measuring insulin action in terms of total body glucose disposal. It is the "gold standard" for quantifying insulin resistance/sensitivity.Diabetic Complications: University of Utah https://www.diacomp.org/shared/document.aspx?id=10&docType=Protocol 2 2019 Donald McClain 2019. Glucose "Clamp" for measurement of whole body insulin-stimulated glucose disposal. protocols.io dx.doi.org/10.17504/protocols.io.7s9hnh6 2021-03-29 03:08:40
Mild Hypothermia to prevent Acute kidney injury in Liver Transplantation (MHALT) Trial - Statistical Analysis Plan for the Interim Analysis, v1.0
 
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Michael Bokoch, Claus Niemann, Dieter Adelmann, Rishi Kothari 10.17504/protocols.io.bf5kjq4w UCSF Transplant Anesthesia Research Group (TARG) Acute kidney injury (AKI), or worsening kidney function, is a common complication after liver transplantation (20-90% in published studies). Patients who experience AKI after liver transplantation have higher mortality, increased graft loss, longer hospital and intensive care unit stays, and more progression to chronic kidney disease compared with those who do not. In this study, half of the participants will have their body temperature cooled to slightly lower than normal (mild hypothermia) for a portion of the liver transplant operation, while the other half will have their body temperature maintained at normal. The study will evaluate if mild hypothermia protects from AKI during liver transplantation.This study is a single-blinded, randomized controlled trial of mild hypothermia during liver transplantation to provide protection from AKI. Participants will be randomized to normothermia (36.5-37.5 °C) versus mild hypothermia (34-35 °C) during a portion of the liver transplant operation. The protocol is based on preliminary data from rodent models showing that hypothermia protects the kidneys from ischemia-reperfusion injury, as well as studies in deceased organ donors showing that cooling improves post-transplant organ function. Temperature will be maintained with standard techniques plus a minimally-invasive esophageal cooling device that is approved by the U.S. Food and Drug Administration. The investigators hypothesize that mild hypothermia will reduce the incidence and severity of AKI after LTx. Standard surrogates (e.g., change in serum creatinine, need for initiation of dialysis) and biomarkers will be used to assess the severity of kidney injury. University of California, San Francisco, University of California, San Francisco, University of California, San Francisco, University of California, San Francisco 1 2020 Michael Bokoch, Claus Niemann, Dieter Adelmann, Rishi Kothari 2020. Mild Hypothermia to prevent Acute kidney injury in Liver Transplantation (MHALT) Trial - Statistical Analysis Plan for the Interim Analysis, v1.0 . protocols.io dx.doi.org/10.17504/protocols.io.bf5kjq4w 2021-03-29 03:08:40
Slide Preparation for Mounting Samples for TissueCyte 1000 Sectioning and Imaging
 
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Allen Institute for Brain Science 10.17504/protocols.io.bdvni65e BICCN, Allen Institute for Brain Science 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. Allen Institute 2 2020 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 2021-03-29 03:08:57
Radioactive in vitro transcription
 
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Anna Behle 10.17504/protocols.io.gudbws6 Axmann Lab Protocol for radioactive labelling of RNA using T7-Polymerase and α-[32P]-UTP. Institute for Synthetic Microbiology, Heinrich-Heine University Düsseldorf 1 2017 Anna Behle 2017. Radioactive in vitro transcription. protocols.io dx.doi.org/10.17504/protocols.io.gudbws6 2021-03-29 03:08:59
Preparing primary T cells for fluorescence microscopy
 
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Bulent Arman Aksoy, Pinar Aksoy, Jeff Hammerbacher 10.17504/protocols.io.vede3a6 Hammer Lab Medical University of South Carolina, Medical University of South Carolina, Medical University of South Carolina 1 2018 Bulent Arman Aksoy, Pinar Aksoy, Jeff Hammerbacher 2018. Preparing primary T cells for fluorescence microscopy. protocols.io dx.doi.org/10.17504/protocols.io.vede3a6 2021-03-29 03:08:58
Stellaris® RNA FISH Simultaneous IF + FISH in Adherent Cells Protocol
 
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LGC Biosearch Technologies 10.17504/protocols.io.ek8bczw Stellaris RNA FISH protocol to simultaneously label adherent cells with IF and RNA FISH. Biosearch Technologies https://biosearchassets.blob.core.windows.net/assets/bti_custom_stellaris_immunofluorescence_protocol.pdf 1 2016 LGC Biosearch Technologies 2016. Stellaris® RNA FISH Simultaneous IF + FISH in Adherent Cells Protocol. protocols.io dx.doi.org/10.17504/protocols.io.ek8bczw 2021-03-29 03:09:00
Protein quantification protocol optimized for zebrafish brain tissue (Bradford method)
 
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Adrieli Sachett, Matheus Gallas-Lopes, Greicy M M Conterato, Radharani , Ana Herrmann, Angelo Piato 10.17504/protocols.io.bjnfkmbn Fish behavior and physiology Zebrafish are incresingly used as a model animal in neuroscience research. Here we describe a protocol to quantify the total amount of proteins in zebrafish brain tissue. Universidade Federal do Rio Grande do Sul, Universidade Federal do Rio Grande do Sul, Universidade Federal de Santa Catarina, Universidade Federal do Rio Grande do Sul, Universidade Federal do Rio Grande do Sul, Universidade Federal do Rio Grande do Sul 1 2020 Adrieli Sachett, Matheus Gallas-Lopes, Greicy M M Conterato, Radharani , Ana Herrmann, Angelo Piato 2020. Protein quantification protocol optimized for zebrafish brain tissue (Bradford method). protocols.io dx.doi.org/10.17504/protocols.io.bjnfkmbn 2021-03-29 03:09:00
Instructions for Planar Area Analysis using CPCe 4.1 Software
 
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Rebecca Maher 10.17504/protocols.io.pmvdk66 Maher RL, Johnston MA, Brandt ME, Smith TB, Correa AMS (2018) Depth and coral cover drive the distribution of a coral macroborer across two reef systems. PLoS ONE 13(6): e0199462. doi: 10.1371/journal.pone.0199462 Oregon State University https://doi.org/10.1371/journal.pone.0199462 1 2018 Rebecca Maher 2018. Instructions for Planar Area Analysis using CPCe 4.1 Software. protocols.io dx.doi.org/10.17504/protocols.io.pmvdk66 2021-03-29 03:08:59
BBM Media
 
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Dr. Steven Wilhelm 10.17504/protocols.io.h97b99n The Aquatic Microbial Ecology Research Group - AMERG (The Buchan, Zinser and Wilhelm labs), CyanoHABs Please contact Dr. Steven Wilhelm ([email protected]) for additional information regarding this protocol.Modified from Bold 1949, Bischoff and Bold 1963 1 2017 Dr. Steven Wilhelm 2017. BBM Media. protocols.io dx.doi.org/10.17504/protocols.io.h97b99n 2021-03-29 03:08:59
In situ hybridization
 
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Michiyo Tsuru 10.17504/protocols.io.j9hcr36 Clinical Proteomics and Gene Therapy Laboratory We generated a miR-340 probe by tagging Has-miR-340 aatcaG(L)t5(L)aT(L)tG(L)cT(L)ttataa_N(6)_Y with Alexa Fluor 488. LNA-in situ hybridization (ISH) was performed according to the manufacturer’s instructions (http://www.exiqon.com/mirna-ish-kit). Sections of FFPE tissues of human OPLL and null mice (4-µm-thick) were fixed with 4 % PFA in PBS for 20 min at room temperature, washed with PBS (3 × 5 min), treated with 0.5 % Triton X-100 (10 min at 4 °С), and briefly washed with PBS followed by two washes (10 min each) in saline-sodium citrate buffer (2× SSC, 0.3 M NaCl, 0.03 M Na3С6Н5О7, рН 7.0). The sections were then digested with 15 μg/mL proteinase K (Exiqon, Vedbaek, Denmark) at 37 °C for 10 min and rinsed for 3 × 5 min in PBS. Hybridization was performed in a humid chamber for 18 h at 65 °C. In situ hybridization of miR-340 was conducted using a miRCURY LNA microRNA ISH Optimization kit (Exiqon, Vedbaek, Denmark). miR-340-Alexa Fluor 488 and bone morphogenetic protein 2 (Alexa Fluor 555) were detected in human OPLL tissue by in situ hybridization and immunocytochemistry, respectively. Nuclei were counterstained with DAPI. After staining, the tissues were observed by fluorescence microscopy (BZ-X700, Keyence, Osaka, Japan). Tsuru M, Ono A, Umeyama H, Takeuchi M, Nagata K (2018) Ubiquitin-dependent proteolysis of CXCL7 leads to posterior longitudinal ligament ossification. PLoS ONE 13(5): e0196204. doi: 10.1371/journal.pone.0196204 Clinical Proteomics and Gene Therapy Laboratory, Kurume University, Fukuoka, Japan https://doi.org/10.1371/journal.pone.0196204 1 2018 Michiyo Tsuru 2018. In situ hybridization. protocols.io dx.doi.org/10.17504/protocols.io.j9hcr36 2021-03-29 03:08:59
DNA clean-up and size selection for long-read sequencing
 
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Ashley Jones, Neeraj Purushotham, Jamila Nasim, Benjamin Schwessinger 10.17504/protocols.io.6kahcse High molecular weight DNA extraction from all kingdoms DNA extractions often contain impurities which limit the output of long-read sequencing technologies. Here a protocol is provided which removes impurities and size selects for longer fragments. To remove residual RNA and protein, an additional RNAse A and Proteinase K treatment is performed. A clean-up with chloroform: isoamyl alcohol (24:1) removes these proteins and other hydrophobic organics such as lipids. A low volume ethanol precipitation and wash is used to concentrate the DNA, hopefully also reducing polysaccharides. Finally, a Short Read Eliminator (SRE) kit by Circulomics is utilised for size selection, which also appears to clean the DNA. This has been trialled for the sorghum rot fungus Macrophomina phaseolina, providing highly promising results with an Oxford Nanopore MinION. One strain yielded 13.71 Gbases with an N50 of 21.75 kb, another strain yielded 9.72 Gbases with an N50 of 43.50 kb. Similar results are expected across many organisms, but have not been tested. Australian National University, University of Southern Queensland, Australian National University, Australian National University 1 2019 Ashley Jones, Neeraj Purushotham, Jamila Nasim, Benjamin Schwessinger 2019. DNA clean-up and size selection for long-read sequencing. protocols.io dx.doi.org/10.17504/protocols.io.6kahcse 2021-03-29 03:08:59
NADH-linked microtiter plate-based assay for measuring Rubisco activity & activation state – PK-LDH
 
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Cristina Rodrigues Gabriel Sales, Anabela Silva, Elizabete Carmo-Silva 10.17504/protocols.io.bf9rjr56 This protocol uses five reactions to couple RuBP carboxylation and 3-PGA formation to NADH oxidation to measure Rubisco activity, based on Scales et al. (2014). Lancaster Environment Centre, Lancaster University, Library Avenue, Lancaster, LA1 4YQ, UK, Biosystems & Integrative Sciences Institute (BioISI), Science Faculty of Lisbon University, Lisbon, 1749-016, Portugal, Lancaster Environment Centre, Lancaster University, Library Avenue, Lancaster, LA1 4YQ, UK 1 2020 Cristina Rodrigues Gabriel Sales, Anabela Silva, Elizabete Carmo-Silva 2020. NADH-linked microtiter plate-based assay for measuring Rubisco activity & activation state – PK-LDH. protocols.io dx.doi.org/10.17504/protocols.io.bf9rjr56 2021-03-29 03:08:57
Salmonella blood culture surveillance: SOP's ITM
 
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Bieke Tack 10.17504/protocols.io.9rah52e Standard operating procedures for Salmonella blood culture surveillance of the unit of tropical bacteriology at the Institute of Tropical Medicine in Antwerp (Belgium):- Blood culture work-up- Pathogen identification- Salmonella serotyping- Antibiotic susceptibility testing Tack B, Phoba M, Barbé B, Kalonji LM, Hardy L, Puyvelde SV, Ingelbeen B, Falay D, Ngbonda D, Sande MABvd, Deborggraeve S, Jacobs J, Lunguya O (2020) Non-typhoidal Salmonella bloodstream infections in Kisantu, DR Congo: Emergence of O5-negative Salmonella Typhimurium and extensive drug resistance. PLoS Negl Trop Dis 14(4): e0008121. doi: 10.1371/journal.pntd.0008121 Unit of Tropical Bacteriology; Department of Clinical Sciences; Institute of Tropical Medicine Antwerp; Belgium https://www.itg.be/E/tropical-laboratory-medicine-unit#/ 1 2019 Bieke Tack 2019. Salmonella blood culture surveillance: SOP's ITM. protocols.io dx.doi.org/10.17504/protocols.io.9rah52e 2021-03-29 03:08:58
CULTURING i3NEURONS (Basic Protocol 6)
 
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Michael S. Fernandopulle, Ryan Prestil, Christopher Grunseich, Chao Wang, Li Gan, Michael E. Ward 10.17504/protocols.io.5jpg4mn Neurodegeneration Method Development Community Following Day 3 replating, i3Neurons should be cultured for at least 1 week before lysis, fixation, or other experimental endpoint, and they remain viable for at least 1 month with proper maintenance. This protocol covers medium conditions, coating of tissue culture dishes with synthetic polymers, recommended plating densities, and maintenance procedures for long-term culture and for specific experimental applications.Cortical Neuron Culture Medium (CM) is sufficient to promote the maturation and longterm maintenance of i3Neurons in culture. i3Neurons express general markers of cortical neurons, as well as specific pre- and postsynaptic markers of glutamatergic excitatory cortical neurons. Since these cells are post-mitotic after 3 days of differentiation (see Basic Protocol 5) and prefer neuron-conditioned medium to fresh medium, maintenance conditions for these cultures are minimal. Generally, half-medium changes every 7 days with fresh, pre-warmed CM are sufficient for culturing beyond d10 (7 days after replating). Neuron attachment and growth also require a strongly adhesive substrate. Coating plates with synthetic polymers such as poly-L-ornithine (PLO) is necessary for i3Neuron attachment, viability, and successful outgrowth. National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland, Gladstone Institute of Neurological Disease, Gladstone Institutes, San Francisco, California, Gladstone Institute of Neurological Disease, Gladstone Institutes, San Francisco, California, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland https://doi.org/10.1002/cpcb.51 1 2019 Michael S. Fernandopulle, Ryan Prestil, Christopher Grunseich, Chao Wang, Li Gan, Michael E. Ward 2019. CULTURING i3NEURONS (Basic Protocol 6). protocols.io dx.doi.org/10.17504/protocols.io.5jpg4mn 2021-03-29 03:08:58

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