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Authors: Hung Liang Pai, Huan Jui Chang

Proper citation: Hung Liang Pai, Huan Jui Chang 2020. Transformation . protocols.io https:// Copy   

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Authors: LI-COR Biosciences
Group: LI-COR Biosciences
Summary: The independent channels of the LI-COR MPX (Multiplex) Blotter make it possible to optimize blocking buffer, primary antibody dilution, and secondary antibody dilution on a single Western blot. Western blotting procedures that generate a blot of 7.0 x 8.5 cm are easily adapted to the MPX format. The process fits into any laboratory’s standard Western blot workflow. Both home- made and pre-cast gels can be used to generate blots. Electrophoresis and transfer to nitrocellulose membrane are performed under standard conditions. Clamping the blot into the MPX Blotter creates up to 24 independent channels, allowing different conditions to be tested in each channel. The range of usable channels per sample is relative to comb size. For Western blot optimization, a single-well gel (“prep gel”) is all that is needed. For this application, any detection method can be used, including near-infrared (NIR) fluorescence and chemiluminescence. This document presents general guidelines for use with the Odyssey® family of Infrared Imaging Systems.

Proper citation: LI-COR Biosciences 2018. One-Blot Western Optimization Using the MPX™ Blotting System. protocols.io https://dx.doi.org/10.17504/protocols.io.grhbv36 Copy   

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Authors: New England Biolabs
Group: New England Biolabs (NEB)
Summary: This is the protocol for dephosphorylation of 5'-ends of DNA using AnP (Antarctic Phosphatase - M0289).

Proper citation: New England Biolabs 2017. Dephosphorylation of 5' -ends of DNA using AnP (M0289). protocols.io https://dx.doi.org/10.17504/protocols.io.irtcd6n Copy   

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Authors: t.a.mcdougall

Proper citation: t.a.mcdougall 2021. Protein Extraction and western blotting. protocols.io https://dx.doi.org/10.17504/protocols.io.bqubmwsn Copy   

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Authors: Victoria Linderberg
Group: Thomas Crouzier Lab
Summary: This protocol is used for investigating biotin-BSM binding to streptavidin on QCM-D sensors.

Proper citation: Victoria Linderberg 2016. Investigating biotin-BSM binding to streptavidin (that in turn is bound to dS-OEG-biotin) on QCM-D sensors. protocols.io https://dx.doi.org/10.17504/protocols.io.egpbbvn Copy   

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Authors: Efthymios Fidanis, Sophia Ward, Laura Cubitt, Amelia Edwards, Debbie Hughes, Michael Hubank, Jerome Nicod
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 transfer of inactivated virus into plates for RNA extraction.Principles of ExaminationThe assay uses the Hamilton robot to transfer 150 μl of 93 samples of inactivated viral media samples from Barcoded 2 ml tubes (Cat no. 72.694.005) to Barcoded 96 well Nunc plates (Cat no. 260251) maintaining sample identity.--The authors wish to thank Heather Ringrose for support with the Hamilton liquid handlingworkstation.

Proper citation: Efthymios Fidanis, Sophia Ward, Laura Cubitt, Amelia Edwards, Debbie Hughes, Michael Hubank, Jerome Nicod 2020. Automated transfer of inactivated virus samples from individual 2ml tubes to 96-well plate. protocols.io https://dx.doi.org/10.17504/protocols.io.bfb2jiqe Copy   

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Authors: Friederike Holderried, Anne Herrmann-Werner
Summary: Transcripts focus group interviews

Proper citation: Friederike Holderried, Anne Herrmann-Werner 2018. Participatory Quality Improvement. protocols.io https://dx.doi.org/10.17504/protocols.io.t5deq26 Copy   

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Authors: yuejun wang
Summary: Teeth were first disinfected with 75% ethanol and then washed with phosphate-buffered saline. Briefly, PDLSCs were separated from periodontal ligament in the middle one-third of the root. Subsequently, MSCs were digested in a solution of 3 mg/mL collagenase type I (Worthington Biochemical Corp., Lakewood, NJ, USA) and 4 mg/mL dispase (Roche Diagnostics Corp., Indianapolis, IN, USA) for 1 h at 37°C. Single-cell suspensions were obtained by cell passage through a 70-μm strainer (Falcon, BD Labware, Franklin Lakes, NJ, USA). MSCs were grown in a humidified, 5% CO2 incubator at 37°C in DMEM alpha modified Eagle’s medium (Invitrogen, Carlsbad, CA, USA), supplemented with 15% fetal bovine serum (FBS; Invitrogen, Carlsbad, CA, USA), 2 mmol/L glutamine, 100 U/mL penicillin and 100 µg/mL streptomycin (Invitrogen, Carlsbad, CA, USA). The culture medium was changed every 3 days. MSCs at passages 3-5 were used in subsequent experiments. Human embryonic kidney 293T cells were maintained in complete DMEM with 10% FBS, 100 U/mL penicillin and 100 µg/mL streptomycin.

Proper citation: yuejun wang 2017. cell cultures. protocols.io https://dx.doi.org/10.17504/protocols.io.iegcbbw Copy   

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Authors: James Thornton Jr
Group: Metafunc Course 2017
Summary: Uses a custom Centrifuge pipeline to assign taxonomy to gene calls. 

Proper citation: James Thornton Jr 2017. Assign taxonomy to gene calls using Centrifuge. protocols.io https://dx.doi.org/10.17504/protocols.io.ksrcwd6 Copy   

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Authors: DNA Pipelines R&D, Peter Ellis, Lesley Shirley
Summary: This SOP describes the procedure for plate based scRNA-seq performed with a commercial available kit from New England BioLabs. Following library construction, samples are pooled in equivolume and quantified, prior to sequencing on the Illumina HiSeq 4000 platform.

Proper citation: DNA Pipelines R&D, Peter Ellis, Lesley Shirley 2020. Commercial automated scRNA-seq workflow. protocols.io https://dx.doi.org/10.17504/protocols.io.bnmvmc66 Copy   

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Authors: Amy Zimmerman, Susanne Wilken
Group: VERVE Net, Worden Lab
Summary: Purpose: To evaluate the influence of nutrient availability on cellular elemental composition (quotas and stoichiometry) in marine picoeukaryotes.Elemental quotas and ratios are assessed under nutrient replete and deplete conditions at the same time to minimize potential variation between experiments. Cells from an exponentially growing culture are concentrated by centrifugation, washed and re-suspended in a small volume of nutrient deplete media. These concentrated cells are used to inoculate triplicate replete and deplete culture flasks at a starting density corresponding to early-exponential growth. Culture growth is monitored daily. When the mean growth rate of the nutrient deplete treatment is half or less of the replete treatment (GRDEP/GRREP

Proper citation: Amy Zimmerman, Susanne Wilken 2017. Nutrient deplete/replete algal culture for elemental analysis. protocols.io https://dx.doi.org/10.17504/protocols.io.gukbwuw Copy   

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Authors: Wen Aw
Group: Cage Studies

Proper citation: Wen Aw 2018. Enzymatic activity of electron transport system Complexes I. protocols.io https://dx.doi.org/10.17504/protocols.io.rq5d5y6 Copy   

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Authors: Moriah Beck, Abby Jurgensmeier
Group: Beck Lab

Proper citation: Moriah Beck, Abby Jurgensmeier 2019. Transformation of Chemically Competent (Smart) Cells. protocols.io https://dx.doi.org/10.17504/protocols.io.ssfeebn Copy   

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Authors: Marda Jorgensen, Jerelyn Nick
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: The purpose of this Standard Operating Procedure (SOP) is to outline procedures for the OCT embedding of HuBMAP fresh frozen specimens.

Proper citation: Marda Jorgensen, Jerelyn Nick 2020. HuBMAP: Embedding Fresh Frozen OCT Samples. protocols.io https://dx.doi.org/10.17504/protocols.io.bcwsixee Copy   

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Authors: Jessica Domingues Lamosa, Lívia R Tomás, Marcos G. Quiles, Luciana R. Londe, Leonardo B L Santos
Summary: This work fo-cused in the problem of Urban mobility, which in an unplanned urban growth sce-nario might generate negative impacts, like traffic jams, air pollution and infras-tructure flaws. Based on real data for the city of São José dos Campos, the mobil-ity of a typical day was represented. These data consist of an Origin-Destinationsurvey: the city was divided into 55 traffic zones and more than 20 thousand peo-ple were asked about the time of departure and arrival of each trip. The devel-opment was divided in 3 steps, pre-processing, processing and post-processing.In preprocessing, an origin destination graph was generated with a 3-dimensionmatrix representation, in language C++, in which each vertex represents a traf-fic zone and the edges are weighted by the flux of people, with 24 time variations,one for each hour of the day. In the processing, in C, the igraph library was usedto calculate the topological properties such as degree (number of connections),clustering coefficient (neighbors redundancy) and diameter (longest distance) ofa network of mobility over a typical day and we also applied the textit walktrapalgorithm for community detection. In the post-processing, using the concept of(geo) graphs, graphs represented with geolocation, the GeoCNet was developed. Itis a tool that allows the creation of a textit shapefile with the topological prop-erties of the graph.

Proper citation: Jessica Domingues Lamosa, Lívia R Tomás, Marcos G. Quiles, Luciana R. Londe, Leonardo B L Santos 2020. Topological indexes and community structure for urban mobility networks: variations in a typical day. protocols.io https://dx.doi.org/10.17504/protocols.io.bjr8km9w Copy   

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Authors: Alexander Niederau, Despoina Trasanidou
Group: iGEM Wageningen 2019

Proper citation: Alexander Niederau, Despoina Trasanidou 2019. FLUORESCENCE LOSS ASSAY. protocols.io https://dx.doi.org/10.17504/protocols.io.7mfhk3n Copy   

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Authors: Jason Kim
Group: Mouse Metabolic Phenotyping Centers
Summary: This experiment involves a spectrophotometric measurement using Roche Cobas Clinical Chemistry Analyzer. Serum levels of aspartate transferase reflect liver function.

Proper citation: Jason Kim 2019. U Mass - Aspartate Transferase. protocols.io https://dx.doi.org/10.17504/protocols.io.xr5fm86 Copy   

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Authors: Leigh Propper, Franchesca Farris, Marda Jorgensen
Group: Human BioMolecular Atlas Program (HuBMAP) Method Development Community
Summary: Poly-Lysine Coverslip Preparation This protocol describes the process of creating Poly-lysine-coated coverslips that are required for mounting tissue sections for the CODEX® experiment workflow.

Proper citation: Leigh Propper, Franchesca Farris, Marda Jorgensen 2020. CODEX - Poly-l-Lysine Cover-Slip Preparation. protocols.io https://dx.doi.org/10.17504/protocols.io.baxyifpw Copy   

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Authors: Saul Moore, Priota Islam
Group: Behavioural Genomics

Proper citation: Saul Moore, Priota Islam 2020. Microbiome Assay 96WP. protocols.io https://dx.doi.org/10.17504/protocols.io.8kbhusn Copy   

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Authors: Bioline
Group: Bioline
Summary: The SensiFAST™ Probe Hi-ROX One-Step Kit has been formulated for highly reproducible first-strand cDNA synthesis and subsequent real-time PCR in a single tube. The kit is formulated for use with probe-detection technology, including TaqMan®, Scorpions® and molecular beacon probes. A combination of the latest advances in buffer chemistry together with a reverse transcriptase and hot-start DNA polymerase system, ensures that SensiFAST Probe Hi-ROX One-Step Kit produces fast, highly-specific and ultra-sensitive one-step real-time RT-PCR. The SensiFAST Probe Hi-ROX One-Step Kit consists of a 2x SensiFAST Probe Hi-ROX One-Step mix, separate reverse transcriptase and RiboSafe RNase Inhibitor.

Proper citation: Bioline 2016. SensiFAST™ Probe Hi-ROX One-Step Kit. protocols.io https://dx.doi.org/10.17504/protocols.io.fyubpww Copy   

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