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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 dx.doi.org/10.17504/protocols.io.nfzdbp6 Copy
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 dx.doi.org/10.17504/protocols.io.bpyfmptn Copy
Authors: Bei Wang, Wen-Hsin Sandy Lee, Cheng-I Wang
Group: Coronavirus Method Development Community
Proper citation: Bei Wang, Wen-Hsin Sandy Lee, Cheng-I Wang 2020. Antibody neutralization assay with SARS-CoV-2 and SARS-CoV pseudovirus. protocols.io dx.doi.org/10.17504/protocols.io.biztkf6n Copy
Authors: Dante Heredia, Thomas Gould
Summary: Protocol for optogenetically stimulating neuronal subtypes in murine colonic myenteric plexus.
Proper citation: Dante Heredia, Thomas Gould 2020. Optogenetically stimulating enteric neurons in the murine large intestine.. protocols.io dx.doi.org/10.17504/protocols.io.bgr9jv96 Copy
Authors: Aaron Bivins, Warish Ahmed, Devin North, Kyle Bibby
Group: Coronavirus Method Development Community, 2019-nCoV Wastewater Epidemiology
Summary: The following protocol describes the GERM Lab workflow for processing wastewater for SARS-CoV-2 via pH adjustment, MCE filtration, extraction, and assay by droplet digital PCR (ddPCR). It is synonymous to method A as described in Ahmed et al. 2020 (the referenced manuscript) with modifications to the consumables and PCR format. It is intended for use in a BSL2+ lab with extra precautions for processing environmental specimens for SARS-CoV-2 as described in the CDC Interim Laboratory Biosafety Guidelines.
Proper citation: Aaron Bivins, Warish Ahmed, Devin North, Kyle Bibby 2020. Wastewater Concentration by Adsorption and Direct Extraction for SARS-CoV-2 RNA Detection and Quantification using RT-ddPCR. protocols.io dx.doi.org/10.17504/protocols.io.bhiuj4ew Copy
Authors: Qiang Wang, Yan Lei, Xiaolan Lu, Guangrong Wang, Qin Du, Xiaolan Guo, Quming Fan, Guoyuan Zhang, Dongsheng Wang
Summary: The serological detection of antibodies to Treponema pallidum is essential to the diagnosis of syphilis. However, for the presence of cross-reaction, the specific antibody tests [e.g., enzyme-linked immunosorbent assay (ELISA)] always have false-positive results. In this study, we derived and validated the dissociation of urea in an attempt to alleviate the situation of false-positive antibodies to T. pallidum detected by ELISA. Six serum samples that were false-positive antibodies to T. pallidumdetected by ELISA, and 16 control serum samples (8 sera positive for both specific IgG and IgM, and 8 IgG-positive and IgM-negative sera) were collected to select the appropriate dissociated concentration and time of urea. Our goal was to establish improved an ELISA method based on the original detection system of ELISA. The sensitivity of the improved ELISA was evaluated by 275 serum samples with class IgM-positive antibodies to T. pallidum. At 6 mol/L with 10 minutes dissociation of urea, 6 samples with false-positive antibodies to T. pallidum were converted to negative, and compared with true-positive antibodies to T. pallidum. The sensitivity of the improved ELISA was 100% by detecting the class IgM-positive antibodies to T. pallidum in sera of patients with syphilis. Considering the importance at the diagnosis of syphilis, antibodies to T. pallidumin serum samples should be retested by the improved ELISA method to avoid false-positive results.
Proper citation: Qiang Wang, Yan Lei, Xiaolan Lu, Guangrong Wang, Qin Du, Xiaolan Guo, Quming Fan, Guoyuan Zhang, Dongsheng Wang 2019. Urea-mediated dissociation alleviate the false-positive Treponema pallidum-specific antibodies detected by ELISA. protocols.io dx.doi.org/10.17504/protocols.io.xdwfi7e Copy
Authors: David Milner
Group: Richards Lab
Summary: Testing for metal toxicity as a result of addition of a putative phosphate transporter to a S. cerevisiae Δpho84 strain
Proper citation: David Milner 2018. S. cerevisiae Δpho84 complementation: metal toxicity. protocols.io dx.doi.org/10.17504/protocols.io.re4d3gw Copy
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 2020. MojoSort™ Human CD14+ Monocyte Isolation Kit Column Protocol. protocols.io dx.doi.org/10.17504/protocols.io.bbwaipae Copy
Authors: Hannigan GD, Grice EA, et al.
Group: VERVE Net, Club Grice
Summary: This protocol provides methods for quality control of metagenomic data. Included is adapter trimming, quality trimming, decontamination, negative control removal, and pre-processing results. Based on the methods found in the following publication:Hannigan, Geoffrey D., et al. "The Human Skin Double-Stranded DNA Virome: Topographical and Temporal Diversity, Genetic Enrichment, and Dynamic Associations with the Host Microbiome." mBio 6.5 (2015): e01578-15.
Proper citation: Hannigan GD, Grice EA, et al. 2016. Script P1: Pre-Processing Samples. protocols.io dx.doi.org/10.17504/protocols.io.edrba56 Copy
Authors: Addgene The Nonprofit Plasmid Repository
Summary: This protocol is for isolating monoclonal cell population by limiting dilution. To see the full abstract and additional resources, please visit the Addgene protocol page.Sample Data1lentiCas9-Blast was a gift from Feng Zhang (Addgene plasmid #52962) and is described in Improved vectors and genome-wide libraries for CRISPR screening. Sanjana NE, Shalem O, Zhang F. Nature Methods. 2014 Aug;11(8):783-4.
Proper citation: Addgene The Nonprofit Plasmid Repository 2019. Isolating a Monoclonal Cell Population by Limiting Dilution. protocols.io dx.doi.org/10.17504/protocols.io.bawwiffe Copy
Authors: Addgene The Nonprofit Plasmid Repository
Summary: The following protocol is for making LB agar plates for the purpose of bacterial selection (500mL of LB agar makes about 25 LB agar plates). Please see the Addgene website for additional details.
Proper citation: Addgene The Nonprofit Plasmid Repository 2019. Pouring LB Agar Plates. protocols.io dx.doi.org/10.17504/protocols.io.5icg4aw Copy
Authors: Caio Maximino, Monica Gomes Lima-Maximino, Diógenes Silva
Summary: Environmental and housing conditions experienced by laboratory animals exert profound effects on their biology, physiology, and behavior. These parameters are important and often overlooked sources of potential variation in experiments, and should be reported in peer-reviewed publications in order to promote scientific reproducibility. To that end, here we provide a summary of the environmental conditions in zebrafish (Danio rerio) at tanks at Laboratório de Neurociências e Comportamento 'Frederico Guilherme Graeff' (LaNeC), located in the Universidade Federal do Sul e Sudeste do Pará, Marabá/PA, Brazil. Included are data on the physico-chemical characteristics of the water, as well health, of zebrafish in separate tanks at LaNeC. This protocol was based on Lawrence's (https://dx.doi.org/10.17504/protocols.io.mrjc54n), and is forked from that.
Proper citation: Caio Maximino, Monica Gomes Lima-Maximino, Diógenes Silva 2019. Zebrafish (Danio rerio) Environmental Summary, LaNeC (Marabá/PA, Brazil). protocols.io dx.doi.org/10.17504/protocols.io.rupd6vn Copy
Authors: Christa Smith
Group: Moran Lab, PriME
Summary: A defined artificial seawater medium for growing bacteria.
Proper citation: Christa Smith 2018. Minimal Basal Medium (liquid). protocols.io dx.doi.org/10.17504/protocols.io.jvdcn26 Copy
Authors: Ji Gao
Summary: Simple measurement of XylE (Catechol 2,3-Dioxygenase) enzyme activity by microplate reader. Catechol 2,3-Dioxygenase can catalyze catechol (1,2-Dihydroxybenzene) to 2-HMS, which has a high absorbance at 377 nm, so we may use photometric mode of microplate reader to measure the concentration of 2-HMS, product of the enzyme.
Proper citation: Ji Gao 2019. Measurement of XylE (Catechol 2,3-Dioxygenase) enzyme activity by microplate reader. protocols.io dx.doi.org/10.17504/protocols.io.7suhnew Copy
Authors: Benoit Besson, Florian Sonthonnax, Magalie Duchateau, Youcef Ben Khalifa, Florence Larrous, Hyeju Eun, Véronique Hourdel, Mariette Matondo, Julia Chamot-Rooke, Regis Grailhe, Hervé Bourhy
Summary: 1st purification: a-FLAG (A2220; Sigma-Aldrich)2nd purification: GFP TRAP_A (ChromoTek)
Proper citation: Benoit Besson, Florian Sonthonnax, Magalie Duchateau, Youcef Ben Khalifa, Florence Larrous, Hyeju Eun, Véronique Hourdel, Mariette Matondo, Julia Chamot-Rooke, Regis Grailhe, Hervé Bourhy 2017. Tandem affinity purification of a bait host protein in presence of a viral protein. protocols.io dx.doi.org/10.17504/protocols.io.jeqcjdw Copy
Authors: David Lowry, Acer VanWallendael
Group: Mimulus
Summary: This is a video of how to plant Mimulus guttatus seeds.
Proper citation: David Lowry, Acer VanWallendael 2019. How to plant Mimulus guttatus. protocols.io dx.doi.org/10.17504/protocols.io.bar8id9w Copy
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 dx.doi.org/10.17504/protocols.io.3qrgmv6 Copy
Authors: Fatma Gomaa, Zhou Zhuha, Roberto Docampo, Virginia Edgcomb, Peter Girguis
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Proper citation: Fatma Gomaa, Zhou Zhuha, Roberto Docampo, Virginia Edgcomb, Peter Girguis 2018. Bodo saltans Cassette for tagging EF1alpha gene__IG BsTub. protocols.io dx.doi.org/10.17504/protocols.io.s5jeg4n Copy
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 dx.doi.org/10.17504/protocols.io.bnkimcue Copy
Authors: Daniel Richter, Parinaz Fozouni, Michael Eisen, Nicole King
Group: Ecology of Marine Plankton (ECOMAP) team - Roscoff, King Lab, Protist Research to Optimize Tools in Genetics (PROT-G)
Summary: The origin of animals, which occurred over 600 million years ago, left no evidence in the fossil record. To trace the earliest events in animal prehistory, we compare extant animals to their closest living relatives, the choanoflagellates, in order to reconstruct the gene content of their last common ancestor and how it evolved on the stem lineage leading to animals. In this project, we increase the accuracy of ancestral animal and choanoflagellate gene content reconstructions by sequencing the transcriptomes of 19 species of choanoflagellates selected for their phylogenetic diversity.
Proper citation: Daniel Richter, Parinaz Fozouni, Michael Eisen, Nicole King 2017. Transcriptome sequencing of 19 diverse species of choanoflagellates. protocols.io dx.doi.org/10.17504/protocols.io.kwscxee Copy
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