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Authors: Nicola Wilson, Fernando Calero-Nieto, Javier Rodríguez-Ubreva, Bertie Gottgens, Esteban Ballestar, Roser Vento-Tormo
Group: Human Cell Atlas Method Development Community, Coronavirus Method Development Community
Summary: This is a protocol for CITE-seq for PBMCs. It includes the steps for Thawing samples and Antibody Staining.

Proper citation: Nicola Wilson, Fernando Calero-Nieto, Javier Rodríguez-Ubreva, Bertie Gottgens, Esteban Ballestar, Roser Vento-Tormo 2020. CITE-seq for PBMCs. protocols.io https://dx.doi.org/10.17504/protocols.io.bjhmkj46 Copy   

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Authors: Ladislav Nevery
Group: Coronavirus Method Development Community, XPRIZE Rapid Covid Testing, Ladislav Nevery
Summary: By passing saliva sample thru 200nm filter to remove particles larger than 120nm covid 19 virus particles and concentrating resulting fluid to very small 1 mm2 area of 100nm filter. Pores in this small area of100nm filter should be clogged up fast raising filter air resistance. Due to high virus density 5.2 log10 ml saliva of infected person should clogg filter much faster than healthy person. Test is setup in such way that act of compressor automatically turning off after reaching threshold PSI value will indicate high count of particles larger than 100nm of being present in measured filter area.

Proper citation: Ladislav Nevery 2020. covid 19 indirect detection thru rise of 100nm filter air resistance . protocols.io https://dx.doi.org/10.17504/protocols.io.bkznkx5e Copy   

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Authors: Josh Quick
Group: Diaz-Munoz Lab, Coronavirus Method Development Community, Szentagothai Research Centre Virology
Summary: This one-pot native barcoding protocol was developed in conjunction with Oxford Nanopore Technologies, New England Biolabs and BCCDC.

Proper citation: Josh Quick 2020. One-pot native barcoding of amplicons v2. protocols.io https://dx.doi.org/10.17504/protocols.io.bdp8i5rw Copy   

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Authors: John Warner, Rebecca Silveston, Delphine Dean
Group: Coronavirus Method Development Community, XPRIZE Rapid Covid Testing, Accessible Health

Proper citation: John Warner, Rebecca Silveston, Delphine Dean 2020. Accessible Health Sentinal Test Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.bk5wky7e Copy   

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Authors: Adélaïde Roguet, Shuchen Feng
Group: Coronavirus Method Development Community, mclellan lab

Proper citation: Adélaïde Roguet, Shuchen Feng 2021. Concentration of viruses from sewage using HA filters. protocols.io https://dx.doi.org/10.17504/protocols.io.bpg7mjzn Copy   

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Authors: Nabil-Fareed Alikhan, Emma Griffiths, Ruth Timme, Duncan MacCannell
Group: Coronavirus Method Development Community, PHA4GE
Summary: PURPOSE: This protocol covers the steps for submitting a SARS-CoV-2 assembly to ENA For new submitters, there's quite a bit of groundwork that needs to be established before a laboratory can start its first data submission. We recommend that one person in the laboratory take a few days to get everything set up in advance of when you expect to do your first data submission. Two protocols cover the PHA4GE guidance for SARS-CoV-2 submission to ENA (Raw sequence data, metadata, and assemblies)Complete in order (1 then 2):1. SARS-CoV-2 EBI submission protocol: ENA, BioSample, and BioProjectStep-by-step instructions for establishing a new Webin laboratory submission account and for creating and linking a new BioProject to an existing umbrella effort.Submit SARS-CoV-2 raw data to ENA (European Nucleotide Archive) and metadata.2. SARS-CoV-2 EBI assembly submission protocol (included protocol) Required: established BioProject and BioSamplesSubmit SARS-CoV-2 assemblies to ENA linking to existing BioProject, BioSamples, and raw data.2. SARS-CoV-2 EBI assembly submission protocol (included protocol) Required: established BioProject and BioSamples

Proper citation: Nabil-Fareed Alikhan, Emma Griffiths, Ruth Timme, Duncan MacCannell 2020. SARS-CoV2 EBI assembly submission protocol . protocols.io https://dx.doi.org/10.17504/protocols.io.bhwqj7dw Copy   

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Authors: Nicola O'Reilly, Svend Kjaer, Maria Greco
Group: Coronavirus Method Development Community, Crick COVID-19 Consortium
Summary: Purpose of examination / Clinical relevanceAt the end of 2019, several pneumonia cases were reported in Wuhan, China and the pathogen was confirmed as a new viral strain. World Health organization has named the newly identified coronavirus as 2019-nCoV, also known as COVID19. The disease developed into a dangerous pandemic, posing major challenges to the NHS. Although more research is necessary to better understand the virus, in response to the emergency, simple and rapid testing is essential to identify the virus in infected individuals. This will aid the implementation of efficient interventions to contain the spread, and distinguish healthcare workers who have been infected, and are required to self-isolate, from those showing similar symptoms but which are not 2019-nCoV associated. The latter category may continue to work, alleviating stress on hard-pressed healthcare resources. 2019-nCoV is an RNA virus, and the diagnostic tests detect viral RNA in swabs from patient airways using a reverse transcriptase PCR assay. Samples are submitted to HSL, an accredited reporting laboratory, and transferred to the Crick for testing.Principles of ExaminationThis procedure involves the preparation of TET buffer for the RNA extraction protocol.

Proper citation: Nicola O'Reilly, Svend Kjaer, Maria Greco 2020. Preparation of TET Buffer. protocols.io https://dx.doi.org/10.17504/protocols.io.bfeujjew Copy   

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Authors: d.noerz
Group: Coronavirus Method Development Community

Proper citation: d.noerz 2021. Modified Zhen et al. SARS-CoV-2 Spike-Gene qRT-PCR assay for highly sensitive detection of the HV69/70 deletion. protocols.io https:// Copy   

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Authors: Brian Glenn St Hilaire, Neva C. Durand, Namita Mitra, Saul Godinez, RM , Alyssa Blackburn, Zane Colaric, Joshua W. M. Theisen, David Weisz, Olga Dudchenko, Andreas Gnirke, Suhas S.P. Rao, Parwinder Kaur, Aviva Presser Aiden, Erez Lieberman Aiden
Group: Coronavirus Method Development Community
Summary: Here, we introduce a low-cost, high-throughput method for diagnosis of SARS-CoV-2 infection, dubbed Pathogen- Oriented Low-Cost Assembly & Re-Sequencing (POLAR), that enhances sensitivity by aiming to amplify the entire SARS-CoV-2 genome rather than targeting particular viral loci, as in typical RT- PCR assays. To achieve this goal, we combine a SARS-CoV-2 enrichment method developed by the ARTIC Network (https://artic.network/) with short-read DNA sequencing and de novo genome assembly. For details on our computational pipeline for automated data processing, including documentation and test set, please visit our Githuh at https://github.com/aidenlab/Polar. .justify:after { content: ""; display:inline-block; width: 100%; }

Proper citation: Brian Glenn St Hilaire, Neva C. Durand, Namita Mitra, Saul Godinez, RM , Alyssa Blackburn, Zane Colaric, Joshua W. M. Theisen, David Weisz, Olga Dudchenko, Andreas Gnirke, Suhas S.P. Rao, Parwinder Kaur, Aviva Presser Aiden, Erez Lieberman Aiden 2020. Pathogen-Oriented Low-cost Assembly & Re-sequencing (POLAR): A highly sensitive and high-throughput SARS-CoV-2 diagnostic based on whole genome sequencing. protocols.io https://dx.doi.org/10.17504/protocols.io.bearjad6 Copy   

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Authors: Ellinor Marving, Alonzo Alfaro-Núñez, Sofie Holdflod Nielsen, Michelle Jørgensen, Anna S. Fomsgaard, Shila Mortensen, Morten Rasmussen, Ria Lassaunière, Charlotte Polacek Strandh, Claus Nielsen, Maiken Worsøe Rosenstierne, Arieh Cohen, Anders Fomsgaard, Katja Spiess
Group: Coronavirus Method Development Community

Proper citation: Ellinor Marving, Alonzo Alfaro-Núñez, Sofie Holdflod Nielsen, Michelle Jørgensen, Anna S. Fomsgaard, Shila Mortensen, Morten Rasmussen, Ria Lassaunière, Charlotte Polacek Strandh, Claus Nielsen, Maiken Worsøe Rosenstierne, Arieh Cohen, Anders Fomsgaard, Katja Spiess 2021. Development of a multiplexed RT-qPCR for the surveillance of SARS-CoV-2 variants of world-wide concern. protocols.io https:// Copy   

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Authors: Tho Ho
Group: Coronavirus Method Development Community, VMMU, XPRIZE Rapid Covid Testing
Summary: This protocol shows procedure for sample handling and detection of SARS-CoV-2 using the AMPHABIO HT-HiThroughput PCR COVID-19 Kit.

Proper citation: Tho Ho 2020. AMPHABIO HT-HiThroughput PCR COVID-19 Kit Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.bqdvms66 Copy   

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Authors: Nabil-Fareed Alikhan, Ruth Timme, Emma Griffiths, Duncan MacCannell
Group: Coronavirus Method Development Community, PHA4GE
Summary: This protocol provides the steps needed to establish a new EBI submission environment for your laboratory, including BioProject(s). Once established, this protocol covers raw read submission to EBI and sample metadata to BioSample.For new submitters, there's quite a bit of groundwork that needs to be established before a laboratory can start its first data submission. We recommend that one person in the laboratory take a few days to get everything set up in advance of when you expect to do your first data submission. Two protocols cover the PHA4GE guidance for SARS-CoV-2 submission to EBIComplete in order (1 then 2):1. SARS-CoV-2 EBI submission protocol: ENA, BioSample, and BioProject (included protocol)Step-by-step instructions for establishing a new EBI (Webin) submission account and for creating and linking a new BioProject to an existing umbrella effort.SARS-CoV-2 raw data submission to ENA (European Nucleotide Archive) and metadata to BioSample.2. SARS-CoV-2 EBI assembly submission protocol Required: established BioProject and BioSamplesSubmit SARS-CoV-2 assemblies to EBI, linking to existing BioProject, BioSamples, and raw data.2. SARS-CoV-2 EBI assembly submission protocol Required: established BioProject and BioSamples

Proper citation: Nabil-Fareed Alikhan, Ruth Timme, Emma Griffiths, Duncan MacCannell 2020. SARS-CoV-2 EBI submission protocol: ENA, BioSample, and BioProject. protocols.io https://dx.doi.org/10.17504/protocols.io.bhwdj7a6 Copy   

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Authors: Qianxin Wu, Chenqu Suo, Tom Brown, Tengyao Wang, Sarah Teichmann, Andrew Bassett
Group: Coronavirus Method Development Community, XPRIZE Rapid Covid Testing

Proper citation: Qianxin Wu, Chenqu Suo, Tom Brown, Tengyao Wang, Sarah Teichmann, Andrew Bassett 2020. INSIGHT: a population scale COVID-19 testing strategy combining point-of-care diagnosis with centralised high-throughput sequencing. protocols.io https:// Copy   

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Authors: Shuchen Feng
Group: Coronavirus Method Development Community, mclellan lab
Summary: Quantification by Droplet Digital PCR (RT-ddPCR)

Proper citation: Shuchen Feng 2021. Quantification by Droplet Digital PCR (ddPCR). protocols.io https://dx.doi.org/10.17504/protocols.io.bpg6mjze Copy   

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Authors: Max Marrone
Group: Coronavirus Method Development Community, Opentrons COVID-19 Testing

Proper citation: Max Marrone 2020. Opentrons COVID-19 testing: Station A, Zymo kit, 24 or 48 samples. protocols.io https://dx.doi.org/10.17504/protocols.io.bezzjf76 Copy   

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Authors: Isabel Ott, Chantal Vogels, Nathan Grubaugh, Anne Wyllie
Group: Coronavirus Method Development Community
Summary: This protocol details recommendations for collecting and processing saliva for SARS-CoV-2 detection, as used in Wyllie et al. 2020, which details results of testing saliva collected from COVID-19+ inpatients and asymptomatic healthcare workers.COVID-19 inpatients. Saliva samples were self-collected by the patient. Upon waking, patients were asked to avoid food, water and brushing of teeth until the sample was collected. Patients were asked to repeatedly spit into a sterile 90 mL specimen collection cup until roughly a quarter full of liquid (excluding bubbles), before securely closing it. For patients unable to provide saliva (such as those on mechanical ventilation), clinical teams were advised that suction could be used to collect saliva into the cup (or a sputum trap container for improved containment and safety). Collected volumes ranged from 0.5 - 20 mL. All saliva samples were stored at room temperature and transported to the research lab at the Yale School of Public Health within 5 hours of collection, with RNA for SARS-CoV-2 detection extracted within 12 hours of collection. When possible, saliva samples were stored at +4°C, otherwise they were kept at room temperature.Asymptomatic healthcare workers. Asymptomatic healthcare workers were asked to collect ~10 mL of saliva into a sterile specimen collection cup. No specific instructions were given regarding food intake etc prior to collection. Samples were delivered to the research lab at the Yale School of Public Health within 6 hours of collection and stored for up to 6 hours at +4°C until aliquoting for RNA extraction. Samples collected on overnight shifts were stored at +4°C before delivery to the research lab.

Proper citation: Isabel Ott, Chantal Vogels, Nathan Grubaugh, Anne Wyllie 2020. Saliva Collection and RNA Extraction for SARS-CoV-2 Detection. protocols.io https://dx.doi.org/10.17504/protocols.io.bg3pjymn Copy   

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Authors: Julia Rossmanith
Group: Coronavirus Method Development Community
Summary: This protocol is for building face shields with closed tops using the print template created by Hanoch Hemmerich on Thingiverse.

Proper citation: Julia Rossmanith 2020. 3D Printed Face Shield with Closed Top. protocols.io https://dx.doi.org/10.17504/protocols.io.bek6jcze Copy   

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Authors: Leigh Jones, Philip Walker, Robert Goldstone, Simon Caidan
Group: Coronavirus Method Development Community, Crick COVID-19 Consortium
Summary: Purpose of examination / Clinical relevanceAt the end of 2019, several pneumonia cases were reported in Wuhan, China and the pathogen was confirmed as a new viral strain. World Health organization has named the newly identified coronavirus as 2019-nCoV, also known as SARS-Cov-2. The disease developed into a dangerous pandemic, posing major challenges to the NHS. Although more research is necessary to better understand the virus, in response to the emergency, simple and rapid testing is essential to identify the virus in infected individuals. This will aid the implementation of efficient interventions to contain the spread, and distinguish healthcare workers who have been infected, and are required to self-isolate, from those showing similar symptoms but which are not 2019-nCoV associated. The latter category may continue to work, alleviating stress on hard-pressed healthcare resources. 2019-nCoV is an RNA virus, and the diagnostic tests detect viral RNA in swabs from patient airways using a reverse transcriptase PCR assay. Samples are submitted to HSL, an accredited reporting laboratory, and transferred to the Crick for testing. The first step of the process is sample receipt at the Crick. This SOP describes the inactivation of the virus.

Proper citation: Leigh Jones, Philip Walker, Robert Goldstone, Simon Caidan 2020. Sample reception, unpacking and barcoding. protocols.io https://dx.doi.org/10.17504/protocols.io.bfe5jjg6 Copy   

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Authors: Chris Ockenhouse, Chris Gast, Renee Holt, Jorge Flores
Group: Coronavirus Method Development Community, PATH
Summary: This is Part 11 of "Phase 3 randomized, double-blinded, placebo-controlled trial to evaluate the safety, immunogenicity, and efficacy of Vaccine Candidate against COVID-19 in adults > 18 years of age"This generic Phase 3 protocol was developed by the PATH team with support of the Bill and Melinda Gates Foundation. The aim of the collection is to share recommended best practices in designing and implementing a Phase 3 study of a COVID-19 vaccine candidate. As Phase 3 trials of different Vaccine Candidates proceed around the world, following the same protocols will ensure consistency and comparability of the Phase 3 trial results.Please note that this is an evolving document, to be versioned and updated, based on community feedback and new data.

Proper citation: Chris Ockenhouse, Chris Gast, Renee Holt, Jorge Flores 2020. Ethical Considerations and Informed Consent (Part 11 of Phase 3 study of Vaccine Candidate for COVID-19). protocols.io https://dx.doi.org/10.17504/protocols.io.bj58kq9w Copy   

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Authors: DNA Pipelines R&D, Benjamin Farr, Diana Rajan, Emma Betteridge, Lesley Shirley, Michael Quail, Naomi Park, Nicholas Redshaw, Iraad Bronner, Louise Aigrain, Scott Goodwin, Scott Thurston, Stefanie Lensing, Carol Scott, Nicholas Salmon, Charlotte Beaver, Rachel Nelson, Alex Alderton, Ian Johnston
Group: Coronavirus Method Development Community
Summary: This SOP describes the procedure for generating cDNA from SARS-CoV-2 viral nucleic acid extracts and subsequently producing 400nt amplicons tiling the viral genome using V3 nCov-2019 primers (ARTIC). This is followed by library construction, equivolume pooling of samples and quantitation, prior to sequencing on the Illumina NovaSeq. It offers the benefit of higher density sample processing in 384 format, whilst matching the data quality achieved in 96 format described in the original protocol:https://www.protocols.io/view/covid-19-artic-v3-illumina-library-construction-an-bgxjjxknBoth the above protocols were adapted from the nCov-2019 sequencing protocol: https://www.protocols.io/view/ncov-2019-sequencing-protocol-bbmuik6w

Proper citation: DNA Pipelines R&D, Benjamin Farr, Diana Rajan, Emma Betteridge, Lesley Shirley, Michael Quail, Naomi Park, Nicholas Redshaw, Iraad Bronner, Louise Aigrain, Scott Goodwin, Scott Thurston, Stefanie Lensing, Carol Scott, Nicholas Salmon, Charlotte Beaver, Rachel Nelson, Alex Alderton, Ian Johnston 2020. COVID-19 ARTIC v3 Illumina library construction and sequencing protocol - high throughput 384 format. protocols.io https://dx.doi.org/10.17504/protocols.io.bnidmca6 Copy   

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