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Authors: Sam Li
Group: BioLegend
Summary: BioLegend MojoSort™ nanobeads work in commonly used separation columns, based on our internal research as well as validation by external testing by academic labs. This simple protocol consists of following the MojoSort™ protocol to label the cells with pre-diluted MojoSort™ reagents and using the columns as indicated by the manufacturer.Note: Due to the properties of our beads, it may be possible to use far fewer beads and less antibody cocktail that with other commercial suppliers. We recommend a titration to find the best dilution factor. However, as a general rule, dilutions ranging from 1:2 to 1:10 for the antibody cocktail can be used. Dilutions ranging from 1:5 to 1:20 for the Streptavidin Nanobeads can be used. Please contact BioLegend Technical Service ([email protected]) if further assistance is needed.
Proper citation: Sam Li 2019. MojoSort™ Mouse Neutrophil Isolation Kit Column Protocol. protocols.io dx.doi.org/10.17504/protocols.io.7bjhikn Copy
Authors: Willem Heijboer, Adam Weir, Eamonn Delahunt, Per Hölmich, Anthony G. Schache, Johannes L. Tol, Robert-Jan de Vos, Zarko Vuckovic, Andreas Serner
Proper citation: Willem Heijboer, Adam Weir, Eamonn Delahunt, Per Hölmich, Anthony G. Schache, Johannes L. Tol, Robert-Jan de Vos, Zarko Vuckovic, Andreas Serner 2020. Adoption and evaluation of the Doha agreement classification system of groin pain in athletes (part 2): a worldwide survey. protocols.io dx.doi.org/10.17504/protocols.io.bgkfjutn Copy
Authors: James Thornton
Group: ECOGEO
Proper citation: James Thornton 2016. Mothur Amplicon Analysis. protocols.io dx.doi.org/10.17504/protocols.io.feibjce Copy
Authors: Janet Keast, Peregrine Osborne
Group: SPARC
Summary: This collection describes the procedures required to label, visualise, characterise and quantify neurons that innervate the lower urinary tract tissues of adult male and female Sprague-Dawley rats. This collection includes protocols for:STAGE 1: Surgery to micro-inject fluorescent retrograde tracer dyes into one or more sites within the lower urinary tractSTAGE 2: Intracardiac perfusion with fixative to preserve neural tissues of interestSTAGE 3: Fluorescence immunohistochemistry of ganglion cryosections.
Proper citation: Janet Keast, Peregrine Osborne 2019. Immunohistochemical classification of sensory and autonomic neurons projecting to the lower urinary tract in rats [keast-001]. protocols.io dx.doi.org/10.17504/protocols.io.w3gfgjw Copy
Authors: Sam Li
Group: BioLegend
Proper citation: Sam Li 2019. Anti-BrdU Staining Protocols Using DNAse with Surface and Fluorescent Proteins. protocols.io dx.doi.org/10.17504/protocols.io.bac9iaz6 Copy
Authors: 宏亮 董
Group: 2019 iGEM NEFU_China
Proper citation: 宏亮 董 2019. Transformation. protocols.io dx.doi.org/10.17504/protocols.io.7idhka6 Copy
Authors: Neilier Junior
Proper citation: Neilier Junior 2020. Enzymatic Assay of Protease Using Azocasein as Substrate. protocols.io dx.doi.org/10.17504/protocols.io.bhqnj5ve Copy
Authors: Maria Victória Magalhães de Vargas, Guilherme Afonso Kessler de Andrade, Sara Navarrete Bohi Goulart, Bruna Mota Bernardes, Filipe Victoria
Summary: Bryophytes are one of the most successful plants in nature and can tolerate several environmental stresses, such as saline stress. MBF1 is a transcription factor that may be involved in reprogramming transcription, which in turn may confer tolerance to stress conditions. The objective of this study was to quantify PaMBF1c under saline stress in three species of moss:Funaria hygrometrica,Physcomitrium acutifolium, andPhyscomitrella patens. All moss species underwent 12 h and 24 h treatment with salt at different concentrations, followed by qRT-PCR experiments.In F. hygrometrica, the MBF1c gene was up-regulated when subjected to 12 h salt treatment at a medium concentration, and may be an important factor for the tolerance of plants to NaCl concentrations. InP. acutifolium,24 h NaCl treatment at of 400, 600, and 800 mM resulted in a gradual up-regulation of MBF1c expression, reflective of the increasing NaCl concentration. InPhyscomitrella,the expression profile of MBF1c showed a down-regulation when treated with 200, 400, and 800 mM NaCl, suggesting that MBF1c may not interfere with the saline stress response in this plant.The diversity of these expression profiles demonstrate that the MBF1c response is not universal neither among plants nor between species of the same family, as shown in the mosses studied.
Proper citation: Maria Victória Magalhães de Vargas, Guilherme Afonso Kessler de Andrade, Sara Navarrete Bohi Goulart, Bruna Mota Bernardes, Filipe Victoria 2020. The role of multiprotein bridging factor 1 (MBF1c) in Funariaceae species under salt stress. protocols.io dx.doi.org/10.17504/protocols.io.be3sjgne Copy
Authors: Andrew Potter
Group: Human Cell Atlas Method Development Community
Summary: Protocol for adult (8-10 week) mouse kidney dissociation performed on ice to reduce artifact gene expression. The first layer, consisting of collagenase digestion, breaks down the tissue and releases some cells and glomeruli and tubules. The second layer consists of bacillus licheniformis digestion for 15 min. augmented with a thermomixer at 1400 RPM and passaging with a 27 gauge needle. The second layer is meant to thoroughly break up remaining tubules and glomeruli, releasing cells such as podocytes. The final yield is 250K cells from 18 mg tissue with 98% viability, approximately 14,000 cells/mg tissue. Approximately 1% of released cells are podocytes (visualized using kidneys from MAFB-GFP+ mice using a hemocytometer).
Proper citation: Andrew Potter 2018. Adult mouse kidney dissociation (on ice). protocols.io dx.doi.org/10.17504/protocols.io.rnmd5c6 Copy
Authors: Josh Quick
Group: Diaz-Munoz Lab, Coronavirus Method Development Community, Szentagothai Research Centre Virology
Proper citation: Josh Quick 2019. Starting a MinION sequencing run using MinKNOW. protocols.io dx.doi.org/10.17504/protocols.io.7q6hmze Copy
Authors: Matthew Sullivan
Group: VERVE Net, Sullivan Lab
Proper citation: Matthew Sullivan 2016. Transcriptomics During One-Step Growth Curves for Cellulophaga Phages. protocols.io dx.doi.org/10.17504/protocols.io.dem3c5 Copy
Authors: Cory Poole, Gloria Pryhuber
Group: Optical Clearing of Tissue
Summary: Purpose and Scope of the Procedure: Provide a relatively rapid clearing of lung tissue sections up to 1 millimeter thick for immunofluorescence staining and imaging with confocal or multiphoton microscopy.Current protocol is limited to tissue sections that are free from severe hemorrhage/thrombosis, containing mostly parenchyma, vessels, and smaller bronchioles. Times and solution volumes may need to be altered slightly if one wishes to clear sections containing a lot of cartilage and/or blood.Scientific Principles: One of the major challenges in conventional brightfield, epifluorescence, and even confocal microscopy is the limitation in imaging depth. This is primarily due to the scattering and absorption of light within the sample of both the excitation light and the emission light leading to a drastic reduction in the image brightness and contrast when deeper than a 100µm. Clearing offers a way around this by both reducing the components that negatively impact the light and by matching the refractive index of the mounting medium to that of the components which make up the tissue, leading to an almost transparent appearance.This protocol is a slight modification of a Clear Unobstructed Brain/Body Imaging Cocktail (CUBIC) protocol described by Kubota et al. developed originally to allow the analysis of cancer metastasis within the bodies of entire mice. By taking advantage of the fact lung tissue has plenty of open spaces and a very large surface area we can reduce the time in each of the solutions to allow complete clearing of the tissue sections to within just a few days.References:·Light Microscopy Methods and Protocols (2017). p. 74·Kubota et al., (2017). Whole-Body Profiling of Cancer Metastasis with Single-Cell Resolution.Cell Reports 20, 236–250
Proper citation: Cory Poole, Gloria Pryhuber 2020. 622.2 URMC HTC Rapid Clearing of Thick Human Lung Tissue Sections. protocols.io dx.doi.org/10.17504/protocols.io.7d7hi9n Copy
Authors: Sebastian Najle
Group: Multicellgenomelab
Summary: This protocol describes the steps for isolation of total RNA for cDNA synthesis in the marine Ichthyosporean Sphaeroforma arctica. In general, Ichthysoporeans are characterized by having a thick cell wall, of uncknown composition, that is particularly hard to brake, making cell lysis difficult. Here we have introduce a simple modification on the conventional RNA extraction protocol using TRIzol reagent, by including two cycles of freezing-tawing of the cells-TRIzol suspension, in order to facilitate cell lysis. This modification improved significantly the RNA isolation yield. Total RNA is further treated with DNAse I enzyme and purified by precipitation to minimize genomic DNA contamination. Highly pure total RNA is then used to synthetize cDNA for downstream applications.
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Proper citation: Sebastian Najle 2019. RNA isolation and cDNA synthesis from the marine ichthyosporean Sphaeroforma arctica. protocols.io dx.doi.org/10.17504/protocols.io.wqdfds6 Copy
Authors: Ken Christensen
Proper citation: Ken Christensen 2020. Western Blot Detection using Licor NIR Fluorescence - CHEM 584 . protocols.io dx.doi.org/10.17504/protocols.io.bj9ckr2w Copy
Authors: Frank Aylward
Proper citation: Frank Aylward 2018. Introduction to Prokaryotic gene prediction (CDS and rRNA). protocols.io dx.doi.org/10.17504/protocols.io.pigdkbw Copy
Authors: Fatma Gomma, Paulo A. Garcia, Jennifer Delaney, Peter R. Girguis, Cullen R. Buie, Virginia P. Edgcomb
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Summary: The process of developing transient transfection protocols in P. caudatus involved initial testing and determination of proper electroporation buffers and parameters (voltage strength, pulse duration and number) to maximize cell viability. Our results suggest that the type of electroporation buffer is critical for maintaining cell viability throughout the experiment and is essential for determining the optimum electric field range.
Proper citation: Fatma Gomma, Paulo A. Garcia, Jennifer Delaney, Peter R. Girguis, Cullen R. Buie, Virginia P. Edgcomb 2017. Parabodo caudatus post-electroporation cell viability quantification using exponential decay electroporation. protocols.io dx.doi.org/10.17504/protocols.io.hmhb436 Copy
Authors: Vance L. Albaugh
Group: Mouse Metabolic Phenotyping Centers
Summary: This is the protocol for conducting the bile diversion procedure in the mouse. In this procedure the common bile duct is ligated proximal to the confluence of the common bile and the pancreatic ducts and an anastomosis is created at a prespecified point along the gastrointestinal tract to re-establish drainage of the biliary tree.
Proper citation: Vance L. Albaugh 2019. Vandy – Bile Diversion in Mice. protocols.io dx.doi.org/10.17504/protocols.io.zdcf22w Copy
Authors: Annette Hayden, Rob Walker, Jack Harrington, Lucy Kimbley, Sara Waise, Chris Hanley, Gareth Thomas, Peter Johnson, Jonathan West, Matthew Rose Zerilli, TIm Underwood
Group: CZI START Project
Summary: A protocol to dissociate fresh esophagus tissue specimens for single-cell transcriptomics.
Proper citation: Annette Hayden, Rob Walker, Jack Harrington, Lucy Kimbley, Sara Waise, Chris Hanley, Gareth Thomas, Peter Johnson, Jonathan West, Matthew Rose Zerilli, TIm Underwood 2021. Single-cell suspensions from primary human esophagus tissue. protocols.io dx.doi.org/10.17504/protocols.io.t9ver66 Copy
Authors: Yoko Kato-Unoki
Proper citation: Yoko Kato-Unoki 2020. ND5 sequencing of Oncorhynchus masou masou. protocols.io dx.doi.org/10.17504/protocols.io.bmf7k3rn Copy
Authors: Angela Piersanti
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Proper citation: Angela Piersanti 2017. Euplotes transfection using Lipofectamine 3000 (provisional). protocols.io dx.doi.org/10.17504/protocols.io.hhub36w Copy
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