eamag/protocols-with-steps huggingface dataset:
TITLE: Method for creating a tissue microarray-capable, slide-scanning acquisition device from any motorized microscope
AUTHORS: Steven P.  Nilsen, M. Lora D. L. M. Ong, Jeremy Muhlich, Jay Copeland, Neal Gliksman, Jerrold R. Turner

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span>High-throughput, multiplex slide-scanning technologies have become widely-used. Advantages include the ability to archive digital copies of slides, review slides as teams using virtual microscope software, and standardize analytical approaches. One barrier to implementation is the cost of dedicated slide-scanning devices. We describe a simple method that allows any microscope to be used for slide-scanning. The only requirements are that the microscope be equipped with a motorized filter turret (for multichannel fluorescence) and a motorized </span><span style = "font-style:italic;">xyz</span><span> stage. This example uses Metamorph, the most commonly used hardware-agnostic microscope control software. The approach can, however, be readily translated to open source applications as well as to dedicated control software provided by microscope manufacturers. The scripts, or journals, developed allow users to define multiple regions of interest and specific parameters, including ranges and channels used for focusing. Following acquisition, captured images can be stitched and displayed using open source virtual microscope software.</span></div></div>

[STEPS]
TITLE: Bogus Data Acquisition Protocol I
AUTHORS: Abby   Moore

[DESCRIPTION]
The purpose of this protocol is to demo protocol development.

The real version of this protocol might be developed with respect to existing publications. Here, I'll refer to publications using the Citation component.


 

 


[BEFORE_START]
This is what you should know before you start.

[GUIDELINES]
These are the individuals responsible for this protocol.

[STEPS]
SECTION: This is my first section
1. 
In the first step, I'll refer to an image of software by using embed code retrieved from Box.

 

SECTION: This is my first section
3. In this step, I'll use the Command component to tell the user to run a command.


 

SECTION: This is my first section
2. In this step, I'll refer to some software by using the Software component.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
Following this protocol, V. Natriegens cells can be transformed with linear fragments and plasmids based on natural transformation as described in Multiplex Genome Editing by Natural Transformation (MuGENT) for Synthetic Biology in Vibrio natriegens  Dalia et. al, 2017. ACS Synthetic Biology.

[STEPS]
SECTION: PRE CULTURE
?. 
SECTION: Transformation reaction
?. 
SECTION: Recovery
?. 
SECTION: Selection
?.
TITLE: NCT04556981 Safety and Immunogenicity of  M72/AS01E in Participants With Well-controlled HIV (MESA-TB)
AUTHORS: Alexander Schmidt; Gates Medical Research Institute

[DESCRIPTION]
Protocol Title: A randomized, placebo-controlled, observer-blind, phase 2 study to evaluate
safety and immunogenicity of the investigational M72/AS01E Mycobacterium tuberculosis (Mtb)
vaccine in virally suppressed, antiretroviral-treated participants with human immunodeficiency
virus (HIV)

Short Title: Safety and immunogenicity of a TB vaccine M72/AS01E in participants with well controlled
HIV

Rationale:
Published phase 1 and 2 randomized, controlled trials evaluating M72/AS01E vaccination in
individuals with HIV who were receiving antiretroviral therapy (ART) show that a 2-dose
schedule of the vaccine given one month apart is well-tolerated and immunogenic in this
population. This current study intends to confirm that the vaccine is safe, well-tolerated, and
immunogenic in a larger population of people with virally suppressed HIV infection in a
tuberculosis (TB) endemic region.

[STEPS]
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>The methods used for re-sequencing the full <em>LPL</em> gene in Kuwaiti Arabs and to analyse the sequence variation and identify variants that could attribute to variation in plasma lipid levels for further genetic association is described in details. Samples (n=100) of an Arab ethnic group from Kuwait were intially selected for the full sequencing of the  30 kb <em>LPL</em> gene locus and flanking regions by Sanger sequencing across the 30 Kb<em> </em>gene and its flanking sequences. A set of 76 primers were designed to cover the entire target sequence yielding on average between 500-700 bp PCR products inlcuding overlapping regions.  The PCR products were then subjected to purification using nucleospin columns.  Each sequence reaction was run in duplicates with one reaction including the forward primer and the other with the reverse primer for quality assurance. Sequence alignment was perfomed to compare the obtained sequences with the genbank reference sequence to screen and identify all varaints.  A selected number of novel variants were selected for validation using realtime PCR.</p>

[STEPS]
TITLE: In vitro alpha-glucosidase enzyme activity
AUTHORS: huseyin akşit

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span style = "font-weight:bold;">alpha-glucosidase enzyme hydrolyses p-NPG to p-nitro phenol and glucoside. p-nitro phenol have max absorbonce at 405 nm.</span></div></div>

[STEPS]
?. Add 50 µl pH=6.8 0.1 M phosphate buffer  each well
50 µl
?. Add 10 µl 1U/ml enzyme solution
10 µl
?. Add 20µl sample solution containing 10, 7.5, 5, 2.5, 1, 0.75, 0.5, 0.25, 0.125  mg/ml perapared in max %20 DMSO:buffer solution
20 µl
37 °C
?. Add 20 µl 0.5 mM p-NPG
20 µl
0 Room temperature
?. Read absorbance at 405 nm
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>Degenerated PCR to test Y-linkage of genes in several Drosophila species. The reactions are made separately for males and females of 400 species and subspecies of Drosophila and related genera.</p>

[STEPS]
SECTION: Pre-Mix Preparation
?. 
SECTION: Final Degenerated PCR preparation
?. 
SECTION: PCR Programs
?.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>Provides a short introduction to MEGAHIT, IDBA-UD, and SPAdes assemblers, a demo on Prodigal Gene Caller, and determining % of reads and contig coverage using Bowtie2 short read aligner.</p>
<p> </p>
<p>Open this protocol inside the virtual machine (details in 'Start Instructions') for easy copy, paste of commands into the command line terminal window. </p>

[BEFORE_START]
<p>Before starting, please visit the ECOGEO website for more information on this "Introduction to Environmental 'Omics" training series. The site contains a pre-packaged virtual machine that can be downloaded and used to run all of the protocols in this protocols.io collection. In addition to the VM, the website contains video and presentations from our initial "Intro to Env 'Omics" workshop held at the Univ. of Hawai'i at Manoa on 25-26 Jul 2016.<br /><br />Please email ‘ecogeo-join@earthcube.org’ to join the ECOGEO listserv for future updates.</p>

[GUIDELINES]
<p><img id="s-mce-img" class="s-mce-img" src="https://s3.amazonaws.com/pr-journal/d9vc2w6.png" data-src="https://s3.amazonaws.com/pr-journal/d9vc2w6.png" data-ofn="Screen Shot 2016-07-12 at 1.52.03 PM.png" /></p>

[STEPS]
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<strong>This general protocol can be used for assessing eukaryotic populations with various diagnostic stains (SYTOX for live/dead cells; DAF-FM for nitric oxide producing cells; DCFDA for reactive oxygen species; others). It is geared for samples that are ~5-10 µm in size, using a 70 µm orifice</strong>.

[STEPS]
TITLE: Coleta de dados actigráficos - ActTrust - V.1
AUTHORS: Daniel Vartanian

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span>Esta coleção reúne os protocolos do processo de coleta de dados actigráficos do Grupo Interdisciplinar de Pesquisa em Sono (GIPSO). O processo foi desenhado para os actígrafos provenientes da companhia </span><a href="https://www.condorinst.com.br/" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">Condor Instruments</span></a><span>. Algumas das configurações mencionadas podem ser específicas desses aparelhos.</span></div><div class = "text-block">Se você não é membro ou parceiro do GIPSO, você pode utilizar esta coleção como referência para o processamento de seus equipamentos. Neste caso, por favor, não envie os formulários mencionados nos protocolos relacionados ao processo.</div><div class = "text-block"><span>Se você utilizar os protocolos do GIPSO em sua pesquisa, por favor considere citá-los na seção de materiais e métodos de sua estudo. Nós nos esforçamos bastante para elaborar e manter protocolos de acesso aberto à comunidade científica. Você pode encontrar a citação dos protocolos na aba</span><span style = "font-weight:bold;"> metadata</span><span>.</span></div><div class = "text-block"><span>Consulte a aba </span><span style = "font-weight:bold;">Guidelines</span><span> para visualizar o fluxograma do processo como um todo.</span></div><div class = "text-block"><span>Para dúvidas ou sugestões, entre em contato pelo e-mail </span><a href="mailto:gipso@usp.br" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">gipso@usp.br</span></a><span> .</span></div></div>

[STEPS]
TITLE: Beetle rearing media
AUTHORS: Yin-Tse  Huang,   Tina

[DESCRIPTION]
ambrosia beetle rearing


[STEPS]
1. The recipe produces enough medium to fill approximately 36 culture tubes.(12cm)
First, add powder or solid, then add liquid
 

2. All ingredients were mixed thoroughly, then put cotton plugs into the mouth of the filled tubes.
3. The culture tubes was autoclaved at 121 °C for 30 min.
4. Put culture tubes to dry 2 night in oven.
(cotton plugs don't remove)
TITLE: Crystallization of Zika virus NS2B-NS3 protease
AUTHORS: xiaomin.ni Ni, Peter Marples, Daren Fearon, Lizbé Koekemoer

[DESCRIPTION]
Zika virus (ZIKV) infections causes microcephaly in new-borns and Guillain-Barre syndrome in adults raising a global public health concern, yet no vaccines or antiviral drugs are available to treat or prevent ZIKV infections. The viral NS3 protease with its NS2B cofactor is essential for the cleavage of Zika polyprotein precursor into individual structural and non-structural proteins and is therefore an attractive drug target. We optimized a robust crystal system of co-expressed NS3 protease with its NS2B cofactor and used this in a crystallographic fragment screening campaign.

[STEPS]
SECTION: Crystallization experiment
2. Protein and buffer requirements:
43.2 µL15 mg/mL 
2.88 mL 
SECTION: Crystallization experiment
4. Dispense 30 µL into SwissCI 3 lens plate reservoir wells using a 100 µl multi-channel pipette.
Dispense 150 nL15 mg/mL  to each lens using the SPT mosquito.
Dispense  150 nL to each lens using the SPT mosquito.

Drop ratio: 1:1
Final drop volume: 300 nl
SECTION: Crystallization experiment
3. Crystallisation screen composition:
  30% w/v PEG 2000
0.2 Molarity (M)  Ammonium sulfate
0.1 Molarity (M)  sodium acetate pH 4.8 

Stock solutions used:
1 Molarity (M)  Ammonium sulfate
1 Molarity (M)  Sodium acetate pH 4.8 
50% w/v PEG 2000

 

SECTION: Crystallization experiment
6. 

 


SECTION: Crystallization experiment
5. Incubate at 20 °C for 1440 min in Formulatrix Rock Imager.

Imaging Schedule: The first images are taken after 12 h and the imaging schedule follows a Fibonacci sequence of days for further collections. 
SECTION: Equipment needed
1. Formulatrix Rock Imager (or incubator of choice)
SPT mosquito
 
P100 8 multi-channel pipette
SwissCI 3 lens plate

SECTION: Data collection at Synchrotron
7. Diamond Light Source
Unattended Data Collection (UDC)
Data Collection Temperature: 100K
Detector: DECTRIS EIGER2 X 9M
Beamline: I04-1
Wavelength:  0.9212 Å
Resolution (Å): 1.62
Beam Size (μm): 60 X 50
Number of images: 3600
Oscillation: 0.10°
Exposure (s): 0.0020
Transmission (%): 100
Flux (ph/s): 9.50e+11
TITLE: Rat brain processing for histological analyses
AUTHORS: miquel.vila

[DESCRIPTION]
Protocol for rat brain processing in order to perform histological analyses

[STEPS]
SECTION: Rat perfusion
1. Deeply anesthetize animals with sodium pentobarbital (50 mg/kg, i.p.)
SECTION: Rat perfusion
2. Perfuse through the left ventricle with saline [0.9% (wt/vol)] at room temperature (RT)
SECTION: Rat perfusion
3. Perfuse again with ice-cold formaldehyde solution 4% in PBS buffered for histology

SECTION: Processing for microtome sectioning
5. Wash twice with 0.1 M PBS and process for paraffin embedding following standard procedures (performed by an external facility) or store brains in 0.1 M PBS at 4 °C, alternatively (not for over a month without changing the PBS)
4. Remove brains and post-fix them for 24 h in the same fixative
SECTION: Processing for cryostat sectioning
6. Cryoprotect for 24-48 h (until they sink) in 30% sucrose at 4 °C
SECTION: Processing for cryostat sectioning
6.1. Exchange sucrose for 0.1 M PBS.
SECTION: Processing for cryostat sectioning
6.2. Immerse brains 30 s in 2-methylbutane and store at -80 °C
SECTION: Sectioning
7. Perform sectioning with a sliding microtome at 5-μm-thickness for paraffin samples or
in a cryostat at 20- or 30-μm-thickness for frozen samples. 
SECTION: Processing for cryostat sectioning
6.3. Include in OCT
TITLE: HEK293 cell culture for co-immunoprecipitation experiments
AUTHORS: Dan Dou, Erika Holzbaur

[DESCRIPTION]
Here, we describe HEK293 cell culture for the purpose of co-immunoprecipitation experiments

[BEFORE_START]
Cells should be routinely tested for mycoplasma contamination using MycoAlert detection kit (Lonza, LT07)

[STEPS]
1. Culture HEK293 cells in DMEM (Corning) supplemented with 10% fetal bovine serum (HyClone)
2. Maintain cells at 37 degrees Celsius with 5% CO2
3. In preparation for co-immunoprecipitation experiments, passage HEK cells with Trypsin onto three 10 cm tissue culture dishes per experimental condition, at a density of 1:10
4. 24 hours later, transfect using FuGENE 6 (6-12 µg total plasmid DNA; Promega)
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>CAR-T therapy has shown significant efficacy in the treatment of leukemia, lymphoma and other blood cancers. Novartis and Gilead Sciences have already received FDA approval for CAR-T therapy targeting the B cell antigen <a href="https://www.creative-biolabs.com/car-t/anti-cd19-car-t-preclinical-in-vivo-assay.htm" target="_blank" rel="noopener noreferrer">CD19</a>. However, these CAR-T therapies often lead to the appearance of side effects of CRS, and even more severe neurotoxicity occurs in some patients. These toxic and side effects have become one of the important reasons limiting the widespread promotion of CAR-T therapy.</p>

[STEPS]
?.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>Zebrafish are an increasingly popular model system to better understand vertebrate physiology. Physical challenge by exercise has become popular to assess physical fitness of genetically manipulated animals or to identify gene regulatory networks that prepare the organism for higher physical demands. The increasing popularity of the zebrafish model system to address questions surrounding physical fitness and physiology has led to the development and commercial availability of sophisticated exercise setups (e.g. swim tunnel respirometers, Loligo® Systems, Viborg, Denmark). However, in many cases, these commercial systems are not available to researchers as they are costly.</p>
<p>This protocol describes a simple and effective way to exercise zebrafish. The setup is assembled from components that are present in virtually any laboratory. Its adoptable to constant or increasing endurance exercise as well as interval exercise or sprint tasks. The procedure does not cause stress to the animals as it relies on their natural behavior to swim against water currents to hold their position relative to their visual environment.</p>

[STEPS]
TITLE: Mitochondrial ROS Determination
AUTHORS: mitsjoecohen  

[DESCRIPTION]
Reactive oxygen species (ROS) is a by-product of biological aerobic metabolism, including oxygen ions, peroxides, and oxygen-containing free radicals. More and more studies have proved that ROS is a messenger molecule of multiple signal pathways, triggered or directly related to cancer, neurodegenerative diseases, fibrosis, cardiovascular and other diseases. Monitoring the level of ROS will help clarify the mechanism of mitochondrial disease and introduce new treatments.

Dichlorofluoresceindiacetate (DCFH-DA) is a ROS probe, which undergoes deacetylation in cells, followed by ROS-mediated oxidation to produce fluorescent substances (excitation wavelength λex=485nm, emission wavelength λem=530nm). DCFH-DA can be used to measure the production of ROS in cytoplasm and organelles (such as mitochondria). The fluorescence intensity can be measured quantitatively by a microplate spectrophotometer.

[STEPS]
TITLE: Love Data Week 2020 at CMU Libraries - Valentine&#39;s Day Dessert Recipes
AUTHORS: Hannah Gunderman, Angelina Spotts, Emma Slayton

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">At CMU Libraries, each year we celebrate Love Data Week (LDW) with events, workshops, speakers, and activities to help engage campus in the world of data that surrounds us! Similar to Open Access Week, the purpose of LDW events is to raise awareness and build a community to engage on topics related to research data management, sharing, preservation, reuse, and library-based research data services. We share practical tips, resources, and stories to help researchers at any stage in their career use good data practices. We believe research data are the foundation of the scholarly record and crucial for advancing our knowledge of the world around us.</div><div class = "text-block">CMU Libraries is proud to offer an institutional license to protocols.io, a collaborative platform for computational workflows, operational procedures, instructions/manuals, and more! It can even be used for cooking recipes! To highlight the research workflow capabilities of protocols.io, we are proud to offer several dessert recipes that are tried-and-true by our 2020 Love Data Week committee at CMU Libraries. Why not make them for a special someone this Valentine's Day?</div><div class = "text-block">We hope you enjoy, and feel free to contact us at UL-DataServices@andrew.cmu.edu if you'd like to use this platform for your research data needs!</div></div>

[STEPS]
TITLE: ERGA_AssemblyProtocol_Template
AUTHORS: ann.mccartney  

[DESCRIPTION]
ERGA_Assembly_Strategy

Abstract written by SAC Committee.

[GUIDELINES]
Any Guidelines Recommended by SAC

[STEPS]
SECTION: Computing Environment Used
1. 
Please enter all details of the compute system your analyses was carried out on

 

SECTION: Data Cleaning
2. Please enter instructions on how your cleaned your data
Software
Summary of approach
Version
Code
System Requirements
Additional Tips for users  
 


SECTION: Assembly Polishing
3. Please enter instructions on how your assembled your long read data
Data Type, Volume  
Summary of approach
Assembly software, Version
 

Purging software
 

System Requirements   
Additional Tips for users
 

Code
 




SECTION: Assembly Scaffolding
4. Please enter instructions on how your scaffolded your long read assembly
Data Type, Volume (before after filtration)  
Summary of approach 
Scaffolding software, Version
 

System Requirements   
Additional Tips for users
 

Code

 

SECTION: Assembly Polishing
5. Please enter instructions on how your polsihed your final assembly
Data Type, Volume  
Summary of approach
Polishing software, Version
 

System Requirements   
Additional Tips for users
 

Code




 



SECTION: Assembly QC
6. QC
Please enter instructions on how your QC'd your final assembly
Summary of approach
Softwares used, Version 
 

GRIT curation employed? 
System Requirements  
Additional Tips for users
 

Code
TITLE: In vitro LRRK2 autophosphorylation
AUTHORS: Xinbo Wang, Pietro De Camilli

[DESCRIPTION]
This protocol details methods for the in vitro LRRK2 autophosphorylation assay.

[STEPS]
SECTION: In vitro LRRK2 autophosphorylation
1. Set up the reaction mixture in a 1.7 mL Eppendorf tube with 1.4 mL purified LRRK2 protein, 1x kinase buffer with 1 millimolar (mM) ATP and 0.01 U/µL GST-Prescission Protease (to remove the Flag tag).
 


SECTION: In vitro LRRK2 autophosphorylation
2. Incubate samples   at 4 °C.
SECTION: In vitro LRRK2 autophosphorylation
3. Add Glutathione beads to remove GST-Prescission Protease.
SECTION: In vitro LRRK2 autophosphorylation
4. Concentrate samples by centrifugal filters and dialyze   at 4 °C against dialysis buffer.
SECTION: In vitro LRRK2 autophosphorylation
5. Check autophosphorylation by Western blotting using a LRRK2 phospho-specific (pT1357) antibody.
SECTION: In vitro LRRK2 autophosphorylation
6. Determine protein concentration by SDS-PAGE using Bovine Serum Albumin (BSA) as standard and used without freezing in the liposome tubulation experiments.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>Purpose of this protocol is to assess the enzymatic activity of glycosidases using fluorophores conjugated to substrates of interest (such as cellobiose or xylobiose). Once the enzyme of interest cleaves substrate, the fluorophore is released enabling quantification of enzymatic activity.</p>

[STEPS]
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?.
TITLE: Immunohistochemistry
AUTHORS: Tiziano Balzano, Javier Blesa, Jose Obeso

[DESCRIPTION]
Immunohistochemistry protocol

[STEPS]
1. Wash sections 3 x 5 min with Tris buffered solution (TB).
2. Antigen retrieval: incubate secions 30 min in citrate buffer at 37°C.
3. Wash sections 3 x 5 min with TB.
4. Wash sections 10 min with Tris buffered-saline solution (TBS).
5. Endogenous peroxidase blocking: incubate sections in a solution consisting of 3% H2O2 + 10% methanol diluted in distilled water for 20 min.
6. Wash sections 3 x 15 min with TBS.
7. Incubation in blocking solution: Incubate sections in a solution consisting of 10% of normal serum (from the same species as the secondary antibody) + 0.3% of Triton X-100 diluted in TBS at RT for 3 hours.
8. Primary antibody incubation: incubate sections 64hr at 4°C with primary antibody diluted in blocking buffer. Negative controls are incubated with blocking buffer alone.
9. Wash sections 4 x 15 min with TBS.
10. Secondary antibody incubation: incubate sections 2hr at RT with biotinylated secondary antibody diluted in blocking buffer. 
11. Wash sections 4 x 15 min with TBS.
12. ABC kit incubation: incubate sections in a solution with avidin-biotin horseradish peroxidase conjugate (ABC) for 45 min at RT.
13. Wash sections 2 x 15 min with TBS.
14. Wash sections 1 x 15 min with TB.
15. Signal was visualized incubating sections in a solution consisting of DAB 0.05% + H2O2 0.003% diluted in TB during 5-10 min under microscope control. 
16. Wash sections with distilled water and counterstain with hematoxylin for 3 min.
17. Wash section with running water for 2 min.
18. Dehydrate sections by immersion in ascending ethanol (5 min each) followed by clearing in xylene for 2 x 10 minutes.
19. Mount the cover slides with DPX.