Protocols   Select Another Resource Report Type

Authors: Andriko Palmowski, Frank Buttgereit

Proper citation: Andriko Palmowski, Frank Buttgereit 2020. Trajectories of Glucocorticoid-Therapy After Initiation in Early (or Methotrexate-Naive) Rheumatoid Arthritis: Protocol for a Systematic Review, Meta-Analysis and Meta-Regression of Observational Cohort Studies. protocols.io https://dx.doi.org/10.17504/protocols.io.bpyfmptn Copy   

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Authors: Marlon Stoeckius, Peter Smibert
Group: Human Cell Atlas Method Development Community
Summary: This protocol is for performing Cell Hashing only. Sample multiplexing and super-loading on single cell RNA-sequencing platforms.Cell Hashing uses a series of oligo-tagged antibodies against ubiquitously expressed surface proteins with different barcodes to uniquely label cells from distinct samples, which can be subsequently pooled in one scRNA-seq run. By sequencing these tags alongside the cellular transcriptome, we can assign each cell to its sample of origin, and robustly identify doublets originating from multiple samples.

Proper citation: Marlon Stoeckius, Peter Smibert 2018. Cell Hashing. protocols.io https://dx.doi.org/10.17504/protocols.io.nfzdbp6 Copy   

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Authors: Curtis Huttenhower
Group: VERVE Net, Huttenhower Lab
Summary: This tutorial focuses on performing a comprehensive metagenomic analysis from whole-genome shotgun sequencing data for microbiome studies. It is divided in to different steps that use the following metagenomic, computational tools: MetaPhlAn, GraPhlAn, LEfSe, and HUMAnN. Our last addition to the PhlAn tools is PhyloPhlAn that can be used in metagenomics for phylgenetically and taxonomically place contigs assembled from whole metagenomic sequencing samples.This tutorial is also a step-by-step description of the metagenomic pipeline we used in our review paper about computational meta'omics (specifically Figure 4).

Proper citation: Curtis Huttenhower 2016. bioBakery Protocols. protocols.io https://dx.doi.org/10.17504/protocols.io.d9899v Copy   

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Authors: Kenny Roberts, Liz Tuck
Summary: For many cutting-edge spatial transcriptomic analysis methods, it is essential to prepare and store tissue in a manner that preserves RNA integrity, tissue morphology, and spatial orientation.This protocol describes the process of freezing fresh or fixed and cryoprotected tissues in optimal cutting temperature compound (OCT) using an isopentane-dry ice slurry. Rapid controlled freezing preserves RNA and mimises ice damage, while embedding in OCT prepares tissues for future cryosectioning.

Proper citation: Kenny Roberts, Liz Tuck 2019. Embedding and freezing fresh human tissue in OCT using isopentane. protocols.io https://dx.doi.org/10.17504/protocols.io.66uhhew Copy   

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Authors: LGC Biosearch Technologies
Summary: Stellaris® RNA FISH protocol for fresh frozen mouse brain

Proper citation: LGC Biosearch Technologies 2016. Stellaris® RNA FISH Protocol for Brain. protocols.io https://dx.doi.org/10.17504/protocols.io.ektbcwn Copy   

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Authors: Kumar Sharma
Group: Diabetic Complications Consortium, Metabolomics Protocols & Workflows
Summary: This protocol describes the procedures for collecting, preparing and performing the DiaComp Creatinine Clearance assay via high performance liquid chromatography (HPLC).Diabetic Complication:

Proper citation: Kumar Sharma 2019. Creatinine Clearance by HPLC. protocols.io https://dx.doi.org/10.17504/protocols.io.8e9hth6 Copy   

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Authors: Laure-Emmanuelle Zaragosi, Pascal Barbry
Group: Human Cell Atlas Method Development Community
Summary: This protocol provides details on the cell dissociation that should be performed to obtain single-cell suspensions from airway biopsies.Biopsies may come from tracheal, bronchial or nasal epithelium.Cell dissociation is performed at 4°C to avoid gene expression alterations and maximize viability. The typical cell number recovery is 40 000 cells for one biopsy.Cell suspensions are suitable for single-cell RNA-sequencing protocols.

Proper citation: Laure-Emmanuelle Zaragosi, Pascal Barbry 2019. Cell dissociation from airway biopsies with cold-active protease for single-cell RNA-seq. protocols.io https://dx.doi.org/10.17504/protocols.io.x3efqje Copy   

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Authors: Isaac Núñez, Daniela Torres, Jens Castor, Aníbal Ignacio Fuentes Palacios, Tamara Matute, Sebastian Rodriguez, Daniel Núñez
Group: RedFungi
Summary: Este protocolo describe los pasos para propagar in vitro hongos tipo terraza (Ver este protocolo para hongos tipo sombrero). Este protocolo ha sido desarrollado bajo la guia de Phil Ross (Mycoworks; https://www.mycoworks.com).

Proper citation: Isaac Núñez, Daniela Torres, Jens Castor, Aníbal Ignacio Fuentes Palacios, Tamara Matute, Sebastian Rodriguez, Daniel Núñez 2018. Cultivo in vitro de hongos tipo terraza. protocols.io https://dx.doi.org/10.17504/protocols.io.ng6dbze Copy   

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Authors: Faezah Mohd Salleh, Jazmin Ramos-Madrigal, Fernando Penaloza, Shanlin Liu, Mikkel-Holger S Sinding, Riddhi P Patel, Renata Martins, Dorina Lenz, Jorns Fickel, Christian Roos, Mohd Shahir Shamsir, Mohammad Shahfiz Azman, Burton K Lim, Stephen J Rossiter, Andreas Wilting, M Thomas P Gilbert
Group: GigaScience Press
Summary: Gen-IALFirst All-tissue DNA extraction kit -This protocol provides an efficient DNA extraction and purification of historic sample (tissue material)

Proper citation: Faezah Mohd Salleh, Jazmin Ramos-Madrigal, Fernando Penaloza, Shanlin Liu, Mikkel-Holger S Sinding, Riddhi P Patel, Renata Martins, Dorina Lenz, Jorns Fickel, Christian Roos, Mohd Shahir Shamsir, Mohammad Shahfiz Azman, Burton K Lim, Stephen J Rossiter, Andreas Wilting, M Thomas P Gilbert 2017. Extraction method F (CR). protocols.io https://dx.doi.org/10.17504/protocols.io.infcdbn Copy   

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Authors: Yaowu Yuan
Group: Mimulus
Summary: This protocol is part of a collection for Mimulus in planta transformation.

Proper citation: Yaowu Yuan 2019. Agro Preparation for Mimulus in Planta Transformation. protocols.io https://dx.doi.org/10.17504/protocols.io.3rqgm5w Copy   

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Authors: Bonnie Hurwitz, Ken Youens-Clark
Group: Hurwitz Lab, Metafunc Course 2017
Summary: This collection of protocols will help you to get started with computational resources needed for the class. We will also get accounts for each of these resources.

Proper citation: Bonnie Hurwitz, Ken Youens-Clark 2017. Week 1: Getting Started. protocols.io https://dx.doi.org/10.17504/protocols.io.jhhcj36 Copy   

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Authors: Linda A. Amaral-Zettler, Markus Bauer, Donna Berg-Lyons, Jason Betley, J. Greg Caporaso, Hugh W. Ducklow, Noah Fierer, Louise Fraser, Jack A. Gilbert, Niall Gormley, James Huntley, Susan M. Huse, Janet K. Jansson, Simon N. Jarman, Rob Knight, Chris L. Lauber, Elizabeth A. McCliment, Sarah M. Owens, Geoff Smith, Luke Thompson, Hege Vestheim, William A. Walters
Group: Earth Microbiome Project
Summary: The 18S protocol detailed here is designed to amplify eukaryotes broadly with a focus on microbial eukaryotic lineages. The primers target the 18S SSU rRNA and are based on those of Amaral-Zettler et al. (2009). The constructs are designed to be used with the Illumina platform.For running these libraries on the MiSeq and HiSeq, please make sure you read the supplementary methods of Caporaso et al. (2012). You will need to make your sample more complex by adding 5-10% PhiX to your run.The outlines of the protocol are the same as the 16S protocol, but different primers, PCR conditions, and sequencing primers are used. In addition, we have designed a blocking primer that reduces the amplification of vertebrate host DNA to be used on host-associated samples, especially those that have a low eukaryotic biomass. Blocking primer strategy is based on Vestheim et al. (2008).

Proper citation: Linda A. Amaral-Zettler, Markus Bauer, Donna Berg-Lyons, Jason Betley, J. Greg Caporaso, Hugh W. Ducklow, Noah Fierer, Louise Fraser, Jack A. Gilbert, Niall Gormley, James Huntley, Susan M. Huse, Janet K. Jansson, Simon N. Jarman, Rob Knight, Chris L. Lauber, Elizabeth A. McCliment, Sarah M. Owens, Geoff Smith, Luke Thompson, Hege Vestheim, William A. Walters 2018. EMP 18S Illumina Amplicon Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.nuvdew6 Copy   

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Authors: Chaoyi Deng, Xiao Wang, Qianmei Zhu, Yanming Kang, Jinlin Yang, Heng Wang
Summary: Backgroud:Since sedation and analgesia are widely accepted by patients and even considered by many gastroenterologists as an integral component of the endoscopic examination, opioids are indispensible in managing pain. Nalbuphine is as effective as morphine as a perioperative analgesic but has not been compared directly with sufentanil in clinical trials.Objectives：The aims of this study were to compare the efficacy and safety of nalbuphine with that of sufentanil in patients undergoing colonoscopy and to determine the optimal doses of nalbuphine in this indicationMethodsTwo hundred and forty consecutive patients aged 18–65 y with an American Society of Anesthesiologists classification of I–II and scheduled for colonoscopy were randomized to receive sufentanil 0.1 µg/kg (group S), nalbuphine 0.1 mg/kg (group N1), nalbuphine 0.15 mg/kg (group N2), or nalbuphine 0.2 mg/kg (group N3).  Baseline vital signs was recorded before the procedure. The four groups were monitored for propofol sedation using the bispectral index and pain relief was assessed using the Behavioral Pain Scale for non-intubated patients. The incidences of respiratory depression during endoscopy, nausea, vomiting, drowsiness, and abdominal distention were recorded in the post anesthesia care unit at 24 and 48 h after colonoscopy.

Proper citation: Chaoyi Deng, Xiao Wang, Qianmei Zhu, Yanming Kang, Jinlin Yang, Heng Wang 2017. Equivalent doses study of nalbuphine and sufentanil for colonoscopy under bispectral index monitoring. protocols.io https://dx.doi.org/10.17504/protocols.io.iq4cdyw Copy   

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Authors: Integrated Islet Distribution Program
Group: Integrated Islet Distribution Program
Summary: To establish a standardized method for the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) sponsored research in the Integrated Islet Distribution Program (IIDP) for preparing a sentinel flask of human islets in order to monitor sterility and islet integrity at the production center. This is a representative sample of the islet preparation that has been distributed to investigators and is held at the distributing center as a quality control sample. The islets are cultured in the same shipping media used for islet shipment, for 18 hours at 4-8°C, simulating the cold shipping conditions followed by 2 days at 37°C culture. The flask of islets are verification in the event of a dispute over sterility or islet quality.

Proper citation: Integrated Islet Distribution Program 2020. Sentinel Flask Preparation for Cold Shipping Protocol of Human Islets. protocols.io https://dx.doi.org/10.17504/protocols.io.bctziwp6 Copy   

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Authors: Dan Needleman
Group: Protist Research to Optimize Tools in Genetics (PROT-G)

Proper citation: Dan Needleman 2017. Protocol for drug sensitivity assay with S. robusta. protocols.io https://dx.doi.org/10.17504/protocols.io.g9ebz3e Copy   

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Authors: Lynn Doran, Steven Burgess
Group: Ag SynBio Lab UIUC

Proper citation: Lynn Doran, Steven Burgess 2021. Preparation of Carbon Sources for Possible Heterotrophic Feedstock for O. tauri. protocols.io https:// Copy   

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Authors: Dr. Steven Wilhelm
Group: The Aquatic Microbial Ecology Research Group - AMERG (The Buchan, Zinser and Wilhelm labs), CyanoHABs
Summary: Please contact Dr. Steven Wilhelm ([email protected]) for additional information regarding this protocol.Adapted from Wetzel and Likens 2000. Limnological Analyses, Springer NY

Proper citation: Dr. Steven Wilhelm 2017. Total Chlorophyll a Measurements by Spectrophotometer. protocols.io https://dx.doi.org/10.17504/protocols.io.ijpccmn Copy   

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Authors: Lixin Wang, Collin Challis, Honghui Liang, Songlin Li, Charless Fowlkes, Aidan Sullivan, Kumar SR, Yvette Taché
Group: SPARC
Summary: Using a multicolor adeno-associate virus system to label the colonic enteric nervous system for digital tracing of individual neurons and nerve fibers in microcircuits in three-dimensions (3D). The methods include viral vectors retro-orbital injection in mice, preparation of colon tissues, microscopy and 3D digital tracing.

Proper citation: Lixin Wang, Collin Challis, Honghui Liang, Songlin Li, Charless Fowlkes, Aidan Sullivan, Kumar SR, Yvette Taché 2020. Multicolor adeno-associate virus labeling and 3D digital tracing of enteric plexus in mouse proximal colon. protocols.io https://dx.doi.org/10.17504/protocols.io.bqavmse6 Copy   

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Authors: Jocelyn Y. Kishi, Sylvain W. Lapan, Brian J Beliveau, Emma R. West, Allen Zhu, Hiroshi M. Sasaki, Sinem Saka, Yu Wang, Constance L Cepko, Peng Yin
Group: Human Cell Atlas Method Development Community

Proper citation: Jocelyn Y. Kishi, Sylvain W. Lapan, Brian J Beliveau, Emma R. West, Allen Zhu, Hiroshi M. Sasaki, Sinem Saka, Yu Wang, Constance L Cepko, Peng Yin 2020. 3: 30mer branch melting temperatures (SABER-FISH). protocols.io https:// Copy   

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Authors: Silvia Domcke, Andrew J. Hill, Riza M. Daza, Cole Trapnell, Darren A. Cusanovich, Jay Shendure
Group: Human Cell Atlas Method Development Community
Summary: We developed an improved assay for single cell profiling of chromatin accessibility that both uses three levels of combinatorial indexing and, in contrast with previous iterations of sci-ATAC-seq and related methods, does not rely on molecularly barcoded Tn5 complexes (sci-ATAC-seq3). Rather, the first two rounds of indexing are achieved by ligation to either end of the conventional, uniformly loaded Tn5 transposase complex (standard Nextera™), while the final round of indexing remains through PCR. Relative to two-level sci-ATAC-seq but similar to sci-RNA-seq3, sci-ATAC-seq3 reduces the per-cell cost of library preparation as well as the rate of collisions, opening the door to experiments on the scale of 10^6 cells. This protocol no longer requires cell sorting, and we also optimized ligase and polymerase choice, kinase concentration, and oligo designs and concentrations, to maximize the number of fragments recovered from each cell. Of note, while maintaining an enrichment in accessible regions, we made the explicit choice to maximize complexity at the expense of specificity for accessible sites. In particular, we found that the fixation conditions could be tuned to adjust the sensitivity (i.e. complexity) vs. specificity (i.e. enrichment in accessible sites) of the assay.

Proper citation: Silvia Domcke, Andrew J. Hill, Riza M. Daza, Cole Trapnell, Darren A. Cusanovich, Jay Shendure 2020. sci-ATAC-seq3. protocols.io https://dx.doi.org/10.17504/protocols.io.be8mjhu6 Copy   

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