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Authors: Bao Thai
Group: Stephen Floor Lab

Proper citation: Bao Thai 2018. WB transfer buffer (1L of 10X). protocols.io https://dx.doi.org/10.17504/protocols.io.mpac5ie Copy   

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Authors: Adéla Krajčová, Tomáš Urban, Petr Waldauf, František Duška

Proper citation: Adéla Krajčová, Tomáš Urban, Petr Waldauf, František Duška 2019. Step-by-step protocol for high resolution respirometry for human heart homogenates. protocols.io https://dx.doi.org/10.17504/protocols.io.3z9gp96 Copy   

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Authors: Ken Lau
Group: BioLegend

Proper citation: Ken Lau 2021. MojoSort™ Whole Blood Human Neutrophil Isolation Kit Protocol. protocols.io https:// Copy   

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Authors: Dana M Cataldo, Andrea B Migliano, Lucio Vinicius
Summary: Training of subjects and execution of flintknapping sessions followed a written protocol strictly followed by a skilled tutor. The general protocol is presented as well as its treatment-specific variants.

Proper citation: Dana M Cataldo, Andrea B Migliano, Lucio Vinicius 2018. Speech, stone tool-making and the two-step evolution of language. Experimental protocols.. protocols.io https://dx.doi.org/10.17504/protocols.io.jyycpxw Copy   

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Authors: Safa Fathy, Hend Mohammed, Asmaa Emad El Din
Summary: Odontogenic lesions are divided into odontogenic cysts and tumors. They affect the osseous marrow and cortex of the jaw bones, and are uniquely derived from the tissues of developing teeth. Odontogenic cysts are cavities lined by odontogenic derived epithelium. They are the most common lesions of the jaw, specially radicular cysts (RC) and odontogenic keratocysts (OKC). While, RC shows an indolent behavior and rarely recurs after surgical removal, the KOT is invasive and destructive. On the other hand, odontogenic tumors are classified according to behavior into benign and malignant in addition, according to origin into epithelial and mesenchymal. The most common benign odontogenic tumor is ameloblastoma.

Proper citation: Safa Fathy, Hend Mohammed, Asmaa Emad El Din 2018. Immunohistochemical expression of Claudin 1 &Claudin 3 in Radicular cyst, Odontogenic keratocyst, Unicystic ameloblastoma and Conventional ameloblastoma. protocols.io https://dx.doi.org/10.17504/protocols.io.t4seqwe Copy   

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Authors: Julie Haendiges, Narjol Gonzalez-Escalona, Ruth Timme, Maria Balkey
Summary: This procedure outlines the protocol for whole genome sequencing of bacterial organisms using the Illumina DNA Prep library preparation kit for sequencing on an Illumina MiSeq sequencer. This document applies to all laboratory personnel in the Division of Microbiology (DM) as well as laboratories in the GenomeTrakr Network.Complete in order:1. DNA Extraction (Manual DNA Extraction or Automated DNA Extraction using the Qiacube)Step-by-step procedures to obtain high quality DNA from isolates in TSB for whole genome sequencing2. DNA QuantitationQuantitation of extracted DNA using the Qubit Flourometer 3. Library Preparation for WGS (Included SOP or Library Preparation using Illumina Nextera XT )Library preparation using NexteraXT or Illumina DNA Prep (previously Nextera DNA Flex)2. DNA QuantitationQuantitation of extracted DNA using the Qubit Flourometer 3. Library Preparation for WGS (Included SOP or Library Preparation using Illumina Nextera XT )Library preparation using NexteraXT or Illumina DNA Prep (previously Nextera DNA Flex)3. Library Preparation for WGS (Included SOP or Library Preparation using Illumina Nextera XT )4. Sequencing using Illumina MiSeq5. Data Quality Checks and NCBI Submission

Proper citation: Julie Haendiges, Narjol Gonzalez-Escalona, Ruth Timme, Maria Balkey 2020. Illumina DNA Prep (M) Tagmentation Library Preparation for use on an Illumina MiSeq Sequencer. protocols.io https://dx.doi.org/10.17504/protocols.io.bcbnisme Copy   

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Authors: Frank Aylward
Summary: This is an example of a simple short read mapping analysis that could be used as part of a transcriptomics or RNA-seq workflow.  Code is intended for use on an Ubuntu 16.04 LTS OS, but it may work on other Unix or Unix-like systems.This tutorial uses data from the following very nice paper from the Hatfull group:Dedrick, Mavrich, Ng, and Hatfull, Expression and evolutionary patterns of mycobacteriophage D29 and its temperate close relatives. BMC Microbiology, 2017. https://doi.org/10.1186/s12866-017-1131-2The tools that are used include:SRA toolkit: https://www.ncbi.nlm.nih.gov/sra/docs/toolkitsoft/bowtie2: http://bowtie-bio.sourceforge.net/bowtie2/index.shtmlsamtools: http://samtools.sourceforge.net/BEDOPS: https://bedops.readthedocs.io/en/latest/content/reference/file-management/conversion/bam2bed.htmlBEDtools: https://bedtools.readthedocs.io/en/latest/Make sure that these tools are installed before starting the tutorial. On a Ubuntu OS you should be able to install most with "sudo apt install", but you may wish to use a package manager such as Anaconda or Miniconda as well. Throughout the tutorial some notes may be made about particular versions of tools. For example, note that the version of SRA toolkit that is installed with apt install often leads to downstream errors, it may be desirable to install this tool with Miniconda instead. Miniconda bash installers can be found here: https://conda.io/miniconda.html

Proper citation: Frank Aylward 2018. Introduction to read mapping for transcriptomics. protocols.io https://dx.doi.org/10.17504/protocols.io.qhedt3e Copy   

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Authors: Max Novelli
Group: SPARC
Summary: Protocol for dataset “RNEL Bladder January 2019”Lower urinary track nerve responses to high-density epidural spinal cord stimulation ( RNEL January 2019 )

Proper citation: Max Novelli 2021. SPARC RNEL Bladder January 2019 protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.xszfnf6 Copy   

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Authors: G-Biosciences
Group: G-Biosciences
Summary: This is a collection of protocols for the G-Biosciences CB-X™ Protein Assay.

Proper citation: G-Biosciences 2016. CB-X™ Protein Assays Collection. protocols.io https://dx.doi.org/10.17504/protocols.io.e5mbg46 Copy   

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Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the process to obtain electrophysiological recordings and cellular contents from neurons in postnatal mouse and/or human brain slices.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.

Proper citation: Allen Institute for Brain Science 2020. Patch-Seq Recording and Extraction. protocols.io https://dx.doi.org/10.17504/protocols.io.bdpei5je Copy   

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Authors: Azeemsha Thacham Poyil, Farshid Amirabdollahian
Summary: This is a protocol for an progressive muscle strength training in a new domain of adaptive robot-assisted training interactions. The protocol was defined based on sports science literature, where the participants would be performing maximum voluntary contraction (MVC) trials at first, then taking a recovery break, then starting with a low-intensity task, and then gradually increasing the difficulty level. The protocol included a preparation stage, followed by initial measurements, a familiarisation session, and finally a performance session. Thirty (17 males, 13 females) healthy participants of at least 18 years old with no history of injury to the upper limb and back were involved in this experiment. Participants were students or staff members of University of Hertfordshire or other volunteers from outside the university. The total duration of the experiment, including the set-up time for each participant, was normally 40-55 minutes. A questionnaire was given as part of the experiment. The participants were asked to fill in a part of the questionnaire at the beginning of the experiment. They were also requested to update the fatigue state in the questionnaire after finishing the experiment.The electromyogram measurements were taken using an EMG acquisition device (g.USBamp amplifier) from g.tec medical engineering GmbH. The data acquisition parameters (sampling rate, channel selection and so on) of the device were configured using Simulink 2017b.Three EMG electrode channels were configured in bipolar mode with a sampling frequency of 1200Hz. Electromyogram (EMG) signals were collected from 3 upper limb muscles (Biceps Brachii, Anterior Deltoid, and Middle Deltoid) of the participants during a robotic interaction. EMG electrodes for Biceps Brachii muscles were connected to Ports 11-12 of g.USBamp amplifier, Frontal Deltoid muscles to Ports 13-14 and Middle Deltoid muscles to Ports 15-16. The experiment set-up included the HapticMaster robotic interface configured for a rowing task. The tasks involved upper limb exercises, which simulated rowing, using a robot arm (HapticMaster robot) as directed by visual instructions on screen and audio cues. In order to support an aesthetically pleasing interactive task for the participants, an animated rowing environment embedded with audio cues and haptic sensation of underwater viscosity were created using the HapticMaster robot. The background on a wide-screen 43 inch LCD monitor would display the front-end of a rowing boat with flowing water, which would potentially motivate the participants for an active involvement in the task. A suitable audio for water flow was played in the background. The HapticMaster robot was programmed to deliver different viscosities under water and above water while rowing. The starting time, the break period and the stopping time of the experiment were guided by audio cues. The task involved moving a robotic end-effector, while an animated boat rowing environment running in front of the participant on an LCD monitor. Kinematic measurements by the robot were logged into separate csv files for each participant.There were be 3 groups of participants in this study. Control 1 Group (Group A) participants did not receive any adaptation from the robot during the interaction and were given break periods at regular intervals. Intervention Group (Group B) interacted with the adaptive robotic environment, which was designed to adjust the difficulty level of the training exercise based on EMG based fatigue indicators. Control 2 Group (Group C) participants had a similar environment as Group B, but the environment only adapted based on the subject-reported fatigue. The participants were asked to continue the exercise until they felt very tired or until they reported fatigue 3 times or until the maximum feasible robotic resistance was reached. Participants were allowed to stop the session in cases discomfort.

Proper citation: Azeemsha Thacham Poyil, Farshid Amirabdollahian 2019. Protocol for Robot-Assisted Progressive Muscle Strength Training. protocols.io https://dx.doi.org/10.17504/protocols.io.5hrg356 Copy   

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Authors: BioLegend, Inc.
Group: BioLegend

Proper citation: BioLegend, Inc. 2017. Immunofluorescence Microscopy Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.hv8b69w Copy   

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Authors: Billie Hunne, Martin Stebbing, Rachel M. McQuade, John B. Furness
Group: SPARC
Summary: Enteroendocrine cells are important regulators of gastrointestinal, digestive and metabolic function. Here we describe protocols for identifying, mapping and characterising these cells in the rat gastric mucosa using immunohistochemistry and high resolution microscopy techniques

Proper citation: Billie Hunne, Martin Stebbing, Rachel M. McQuade, John B. Furness 2019. Immunohistochemistry and high resolution microscopy of rat gastric enteroendocrine cells. protocols.io https://dx.doi.org/10.17504/protocols.io.xeyfjfw Copy   

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Authors: Atsuko Mizoguchi
Summary: Total RNA was extracted from cells and the cultured supernatant and exosome which was extracted from the same amount of supernatant using 3D-Gene® RNA extraction reagent.Comprehensive miRNA and mRNA expression analysis were performed using 3D-Gene® Human miRNA Oligo Chip (miRBase ver.21) and mRNA Oligo Chip(Toray Industries, Inc.) which are featured with the columnar structure and bead-mixing for high sensitivity.

Proper citation: Atsuko Mizoguchi 2018. Purification of total RNA (microRNA and mRNA) from liquid samples. protocols.io https://dx.doi.org/10.17504/protocols.io.vu3e6yn Copy   

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Authors: Lgc Biosearch Technologies
Summary: A set of Stellaris FISH Probes comprises up to 48 singly labeled oligonucleotides designed to selectively bind to targeted transcripts. Stellaris FISH Probes bound to target RNA produce fluorescent signals that permit detection of single RNA molecules as diffraction-limited spots by conventional fluorescence microscopy. Please see the Biosearch

Proper citation: Lgc Biosearch Technologies 2016. Stellaris® RNA FISH Protocol for Frozen Tissue. protocols.io https://dx.doi.org/10.17504/protocols.io.cdqs5v Copy   

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Authors: Miriam Heinen
Summary: This staining was performed to detect fetuin-A in activated microglia. CD68 is a marker, that stains activated microglia, macrophages and monocytes. Fetuin-A and microglia were detected in paraffin sections (1 μm thickness) of formalin-fixed human brain tissue. CD68 was stained using a monoclonal mouse-anti-human antibody (Dako Cat# M0814, RRID:AB_2314148, clone KP1, dilution 1:50) and a polyclonal goat-anti-mouse Alexa Fluor® 488 conjugated secondary antibody (Thermo Fisher Scientific Cat# A-11029, RRID:AB_2534088, dilution 1:300). Fetuin-A was detected by using a monoclonal IgG2a mouse-anti-human antibody (clone MAHS-1, dilution 1.0 µg/mL), raised against purified human fetuin-A in our laboratories. Antibody binding was detected by tyramide signal amplification using a secondary biotinylated polyclonal goat-anti-mouse antibody (Dako Cat# E0433, RRID:AB_2687905, dilution 1:300) and a Tyramide Signal Amplification Kit (Life Technologies, Carlsbad, USA, T-20933). To minimize lipofuscin autofluorescence, sections were counterstained with Sudan Black (Sigma-Aldrich, Munich, Germany, 199664, dilution 0.3% in 70% ethanol, 5 minutes). Nuclei were stained with DAPI (Sigma-Aldrich, Munich, Germany D9542, dilution 0.25 µg/ml, 5 minutes). Sections were mounted with Immumount (Thermo Scientific, Waltham, USA, 9990402) and stored at 8°C in the dark. 

Proper citation: Miriam Heinen 2018. Immunofluorescence staining protocol for co-staining of fetuin-A and CD68 in older human autopsy tissue. protocols.io https://dx.doi.org/10.17504/protocols.io.syfeftn Copy   

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Authors: Noriko Otsuki
Summary: Functional SMN protein of peripheral blood-derived mononuclear cells was detected with an anti-SMN antibody labeled with a fluorescent dye and analyzed semiquantitatively using intracellular expression intensity and SMN spot formed in cell nucleus as an index, Consider the relationship with the clinical condition and motor function.Briefly,1) In order to identify the cell fraction, the fluorescently labeled cell surface antigen-specific antibody is added to the whole blood sample and stained.2) Hemolyze peripheral blood and fix mononuclear cells.3) Intracellular staining is performed with SMN protein and nuclear-specific fluorescently labeled antibody.4) SMN protein expression analysis was performed on a strongly CD33 cell population detected as a cell surface marker, SMN spot detection algorithm was used to analyze the proportion of cells in which intracellular SMN expression and SMN protein aggregated in the nucleus.

Proper citation: Noriko Otsuki 2018. Staining of survival motor neuron (SMN) protein in peripheral blood mononuclear cells (PBMC) protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.qqkdvuw Copy   

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Authors: Eva Feldman
Group: Diabetic Complications Consortium
Summary: Summary:Oxidative stress is highly correlated with the metabolic changes caused by hyperglycemia. Increased levels of glucose overload mitochondria and result in the production of reactive oxygen species (ROS). In addition, the flow of excess glucose through cellular pathways decreases the cell’s normal ability to detoxify ROS. As a result, the neurons and axons of the peripheral nervous system contain increased levels of ROS and decreased antioxidant capacity. The following assays are used to measure these changes in rodent models of diabetic neuropathy. Diabetic Complication:

Proper citation: Eva Feldman 2019. Biochemical Measures of Neuropathy - Glutathione Peroxidase. protocols.io https://dx.doi.org/10.17504/protocols.io.3qrgmv6 Copy   

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Authors: Megan Folkerts, mnguyen , Danielle Vazquez, Amber Jones
Group: Coronavirus Method Development Community

Proper citation: Megan Folkerts, mnguyen , Danielle Vazquez, Amber Jones 2020. High-Throughput Tiled Amplicon Sequencing of Sars-CoV2 using Seqwell's plexWell 384. protocols.io https://dx.doi.org/10.17504/protocols.io.bnkimcue Copy   

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Authors: Laise Moraes
Summary: The purpose of this protocol is to define and provide instructions for creating a conda environment for phylogenetic analysis

Proper citation: Laise Moraes 2020. Installation instructions for RNA-seq analysis using a conda environment. protocols.io https://dx.doi.org/10.17504/protocols.io.bgffjtjn Copy   

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