eamag/protocols-with-steps huggingface dataset:
TITLE:        ELISA for quantification of IL-41 in human serum or plasma.
AUTHORS: Angel  Justiz-Vaillant

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span>Interleukins (IL) are a type of cytokine first thought to be expressed by leukocytes alone but have later been found to be produced by many other body cells. They play essential roles in the activation and differentiation of immune cells, as well as proliferation, maturation, migration, and adhesion. They also have pro-inflammatory and anti-inflammatory properties. The primary function of interleukins is, therefore, to modulate growth, differentiation, and activation during inflammatory and immune responses. Interleukins consist of a large group of proteins that can elicit many reactions in cells and tissues by binding to high-affinity receptors in cell surfaces.  </span><a href="https://www.ncbi.nlm.nih.gov/books/NBK499840" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">[1]</span></a></div><div class = "text-block">The immunoregulatory cytokine IL-41 (also known as meteorin-like protein) is expressed at high levels in the synovium of patients with psoriatic arthritis (PsA).[2]</div><div class = "text-block">Reference</div><div class = "text-block"><span>1. Justiz Vaillant AA, Qurie A. Interleukin. In:</span><span style = "font-style:italic;">StatPearls</span><span>. Treasure Island (FL): StatPearls Publishing; June 12, 2019.</span></div><div class = "text-block"><span>2.Onuora S. Novel cytokine, IL-41, linked with PsA.</span><span style = "font-style:italic;">Nat Rev Rheumatol</span><span>. 2019;15(11):636. doi:10.1038/s41584-019-0314-7 </span></div></div>

[STEPS]
?. An anti-human IL-41 coating antibody is adsorbed onto the microwells by incubation overnight at 4°C with carbonate-bicarbonate buffer.
?. Add 50 µl of human serum. Human IL-41 present in the serum sample binds to antibodies adsorbed into the microwells.
?. The microplate is blocked with 3% non-fat milk-PBS buffer and later wash to remove unbound proteins.
?. Fifty (50) µl of biotin-conjugated anti-human IL-41 antibody is added. The optimal dilution must be investigated.
?. The microplate is rewashed with PBS-Tween 20 buffer, pH 7.4.
?. One hundred µl of streptavidin-HRP conjugate is added and it binds to the biotin-conjugated anti-human IL-41 antibody. The optimal dilution of this conjugate must be investigated.
?. The plate is washed following incubation to remove the unbound Streptavidin-HRP.
?. Add 100 µl of 3',3',5',5'- tetramethylbenzidine (TMB; Sigma-Aldrich) into each well.
?. Incubate the microwells in the dark for 15 min.
?. A colored product is formed in proportion to the quantity of human IL-41 present in the sample or standard.
?. The reaction is terminated by addition of 100 µl 3M H2SO4  and the absorbance is measured at 450 nm.
?. A standard curve is made from 7 human IL-41 standard dilutions and the human IL-41 sample concentration is determined.
?. For better results place the microplate on a microplate shaker in every incubation.
TITLE: Chemically Competent Cell Preparation [Calcium Chloride Method]
AUTHORS: Harry  Akligoh, Jenny  Molloy

[DESCRIPTION]
This protocol is a low-cost approach for preparing competent cells using 0.1M CaCl2 solution which is useful in cases where commercial competent cells are unaffordable, it is challenging to receive products on dry ice or you do not have an ultra low temperature freezer.

[BEFORE_START]
Disinfect your bench and pipettes using bleach or 70% isopropanol solution 

[GUIDELINES]
Maintain aseptic technique at all point during this experiment

[STEPS]
SECTION: Overnight plating of E. coli
1. Plate an innoculum of Escherichia coli  (e.g. BL21(DE3), JM109) glycerol stock on LB agar without antibiotics.
 

 

 


  
SECTION: Overnight culture
3. Pick and transfer a discrete single colony of E. coli into 10 ml LB broth using a sterile innoculation loop or toothpick.
SECTION: Harvesting cells and Washing of Cells
5. Sub-culture the overnight culture by adding 500 µL into 50 mL  LB broth (i.e. a 1:100 diution) and shake at    

SECTION: Harvesting cells and Washing of Cells
7.  Keep the culture on ice  and only mix gently while harvesting the cells, avoid vigorous shaking.

SECTION: Harvesting cells and Washing of Cells
7.1. Pipette 1 mL culture into a 1.5 mL  microcentrifuge tube and centrifuge at  . 
 


SECTION: Harvesting cells and Washing of Cells
8. After each centrifugation, discard the supernatant making sure not to dislodge the cell pellets and repeat previous step as needed to harvest more cells. If using 1.5 mL tubes, we recommend repeating 3-5 times.

SECTION: Harvesting cells and Washing of Cells
9. Resuspend the harvested cells in 1 mL ice cold 0.1 Molarity (M) calcium chloride (CaCl2) solution.

SECTION: Harvesting cells and Washing of Cells
10. Incubate on ice  for 30 min 

SECTION: Harvesting cells and Washing of Cells
11. Centrifuge to pellet the cells at   

SECTION: Harvesting cells and Washing of Cells
12. Finally, gently resuspend the cells in 500 µL  ice cold 0.1 Molarity (M) calcium chloride (CaCl2) solution.

 


SECTION: Harvesting cells and Washing of Cells
13. Pipette 50 µL  into sterile, ice-cold 1.5 mL  microcentrifuge tubes to use immediately for transformation or for storage at -80 °C .

 

SECTION: Overnight culture
4.  Incubate the culture overnight in a shaker incubator at  

SECTION: Overnight plating of E. coli
2. Incubate the plate37 °C 120 min 
 

SECTION: Harvesting cells and Washing of Cells
6. Monitor OD600 every 30 min using a photometer or McFarland Standards until OD600 = 0.3
TITLE: Determination of phenolic compounds
AUTHORS: Jorge Carlos Ruiz Ruiz

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">Phenolic compounds determined using the Folin-Ciocalteu reagent.</div></div>

[STEPS]
?. An aliquot of sample (0.25 mL) was mixed with 0.15 mL of the Folin-Ciocalteu reagent and 1 mL of a 15% sodium carbonate solution.
?. Distilled water was added to a final volume of 2.5 mL. Following incubation for 3 min at 37 °C in a water bath, the absorbance was measured at 750 nm. Gallic acid (Sigma-Aldrich) was used as the standard and the results were expressed as µg/mL of gallic acid equivalents (GAE).
?. Following incubation for 3 min at 37 °C in a water bath, the absorbance was measured at 750 nm.
?. Gallic acid was used as the standard and the results were expressed as µg/mL of gallic acid equivalents (GAE).
TITLE: 10x Protocols: Visium Fresh Frozen Library Construction -- University of Minnesota TMCs (CG000239 Rev F)
AUTHORS: 10x Genomics, Laura J Niedernhofer, David A Bernlohr

[DESCRIPTION]
Protocols from 10x Genomics for Visium Spatial Gene Expression on Fresh Frozen / OCT samples. 
Completed without CytAssist component.

Version of protocol used: 
   10x protocol CG000239, Revision F -- Library Construction

Additional protocols added for guidance:
    10x protocol for imaging guidelines (CG000241, Rev D)
    10x protocol for tissue preparation guidelines (CG000240, Rev D)

Note: These protocols may not be the current version offered by 10x Genomics but were used to produce the specific datasets connected to them. Please review the 10x Genomics support website for the most recent versions of the protocols prior to starting your experiment. 

[STEPS]
1. 10x protocol CG000239, Rev F (Library construction):
  

2. Additional Protocols/Guidelines
  CG000241, Rev D
  CG000240, Rev D
https://www.10xgenomics.com/support/spatial-gene-expression-fresh-frozen
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p><em>This</em> s<em>ampling protocol is designed to collect two fractions: cell pellets and cell-free filtered supernatants. The cell pellets are used for gene expression analyses by RNA-Seq and proteomics. The cell-free fraction is used for analysis of dissolved analytes (e.g. N, P, DOC). </em></p>
<p><em>Volumes are based on our experience with cyanobacterial cultures at cell densities ~ 5E7/ml. For more/less concentrated cultures, adjust volumes accordingly. </em></p>

[STEPS]
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TITLE: Iris Classification
AUTHORS: Ziyong Ma

[DESCRIPTION]
It is a simple guide for iris classification

[STEPS]
SECTION: Running Iris Classification Project
1. Access the capsule for this project at https://codeocean.com/capsule/0923404/tree.
This project is to build different models to identify different species of flowers.
SECTION: Running Iris Classification Project
2. The data which are images of iris are stored in the images files of data.
SECTION: Running Iris Classification Project
3. Click the "Reproducible Run" to generate a html.file for the ipynb file which contains the code.
TITLE: Isolation of Nuclei from Adult Human Brain Tissue for 10x Genomics Platform
AUTHORS: Allen Institute for Brain Science

[DESCRIPTION]
Isolation of nuclei from frozen adult human brain tissue or thawed and microdissected brain tissue sections for RNA-seq analysis. 

Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U01MH114812. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

[STEPS]
TITLE: ASAP protocol: Crystallization of Enterovirus A71 3C protease (PDB code 8CNY)
AUTHORS: ryan.lithgo, Peter Marples, daren.fearon, Lizbé Koekemoer

[DESCRIPTION]
The crystallization protocol and buffer conditions used to obtain Enterovirus A71 3C protease crystals. The crystal structure was depostited to RCSB PDB with code 8CNY.

[STEPS]
SECTION: Crystallization experiment
2. Protein and buffer requirements:
43.2 µL1 millimolar (mM) 
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 nL1 millimolar (mM)  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:
0.1 M Tris-HCl (pH 8.50)
0.2 M Magnesium chloride hexahydrate 
30% w/v PEG 4000

Stock solutions used:
1 M Tris-HCl adjusted to pH 8.5 with HCl
1 M MgCl2.6H20
50% w/v PEG 4000

 

SECTION: Crystallization experiment
6. Crystal typically form after ~24hrs
 


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

Imaging Schedule: The first images are taken after 12hrs 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 multi-channel pipette
SwissCI 3 lens plate
TITLE: Arabidopsis growth medium with varying sulfur and nitrogen content
AUTHORS: Meike Burow

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">Media receipe to grow A. thaliana seedlings on plates. </div><div class = "text-block">Variaiton in S and N concentrations.</div></div>

[STEPS]
?. [for 1 L medium]
AB1General media recipeVolume2Minor salts (100x)10 mL3Major salts (10x)100 mL4Ferrous solution (100x)10 mL5Vitamin mix (1000x, Murashige and Skoog, Prod. No. M0409.0100, Duchefa Biochemie)1 mL67water to 1L8adjust   pH to 5.8 with 1M KOH9divide   1L to 2x 500 mL (into 1L bottles)10add   4g agar per 500mL11autoclave12
AB1General media recipeVolume2Minor salts (100x)10 mL3Major salts (10x)100 mL4Ferrous solution (100x)10 mL5Vitamin mix (1000x, Murashige and Skoog, Prod. No. M0409.0100, Duchefa Biochemie)1 mL67water to 1L8adjust   pH to 5.8 with 1M KOH9divide   1L to 2x 500 mL (into 1L bottles)10add   4g agar per 500mL11autoclave12
?. [Concentrations of S and N in final medium]
A1Concentration of Sulphate2in +S: 1.68 mM3in -S: 0.015 mM45Concentration of Nitrate6in +N: 3 mM7in -N: 0.3 mM
A1Concentration of Sulphate2in +S: 1.68 mM3in -S: 0.015 mM45Concentration of Nitrate6in +N: 3 mM7in -N: 0.3 mM
?. [stock solutions]
AB1Media
  solutionstock solutions&nbsp;2+S/+NA+C+G3+S/-NA+D+G4-S/+NB+E+H5-S/-NB+F+H
AB1Media
  solutionstock solutions&nbsp;2+S/+NA+C+G3+S/-NA+D+G4-S/+NB+E+H5-S/-NB+F+H
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
This protocol is intended to help you use HOBO data loggers and HOBOware software.

[STEPS]
SECTION: HOBOware
?. 
SECTION: Connection
?. 
SECTION: HOBOware Basic functions
?. 
SECTION: HOBO data logger Programing
?. 
SECTION: HOBO data logger Reading and Data export
?.
TITLE: Set Up Biodata Resource Inventory in Google Colab
AUTHORS: Kenneth  Schackart

[DESCRIPTION]
This protocol will guide you on how to get everything in place to update the Biodata Resource Inventory.

This protocol describes how to setup Google Colab, connect your Google Drive, and clone the repository.

Some of the information here came from the following article: https://medium.com/analytics-vidhya/how-to-use-google-colab-with-github-via-google-drive-68efb23a42d

[STEPS]
SECTION: Prepare Google Drive
1. In your Google Drive home directory, create a new folder called GitHub.

Inside the GitHub folder, create another folder called biodata_resource_inventory.
SECTION: Connect Colab to Drive
2. Go to Google Colab.

If you need to change the account you are using, close the pop up by clicking Cancel at the bottom right. Change your Google account by clicking the icon at the top right.
SECTION: Connect Colab to Drive
3. A menu will pop up. At the bottom right, select: "New Notebook"
SECTION: Connect Colab to Drive
4. In the cell, paste or type the following:

 
Click the "run" button to the left of the cell or run with ctrl+enter

You will be prompted to allow Google Colab to access your Drive, say yes.
 

SECTION: Connect Colab to Drive
5. Create a new code chunk by clicking "+ Code" under the top-left banner. In that cell paste or type the following, then run:

 

This has changed your current directory to biodata_resource_inventory.
 

SECTION: Create GitHub Access Token
6. Go to GitHub.

Go to Settings, navigate to Developer settings and click on Personal access tokens.
SECTION: Create GitHub Access Token
7. Click Generate new token button on the top right corner of the page.

SECTION: Create GitHub Access Token
8. Click the repo checkbox under Select scopes.
SECTION: Create GitHub Access Token
9. Click the Generate token button at the bottom of the page.
SECTION: Create GitHub Access Token
10. You have now created a new access token. Copy the token to your clipboard. Do not close this page yet.

 

SECTION: Clone the Repository
11. Go back to your Colab Notebook, create a new Code cell, and enter the following, pasting your access token into the quotations, then run:

 

SECTION: Clone the Repository
12. Create a new code cell and paste or type the following, then run:
 

Note: During development, the main branch is mostly empty. To switch to the development branch run the following in a code cell:

 

Congrats! You now have linked Google Colab to GitHub for access to their API, and you have cloned the repository into your Google Drive. You can now close the page with your access token.
SECTION: Running the pipelines
13. You can now access all the files in the repository via traditional Google Drive.
SECTION: Running the pipelines
14. You will find two notebooks in the repository:
running_pipeline.ipynb: This allows reproduction of the original results
updating_inventory.ipynb: This guides you to update the inventory

Each notebook provides instructions on their respective process, and includes already filled-in code chunks. So you should have to do little more work to run those pipelines.
TITLE: Peptostreptococcal protein L and a Chimeric Protein-LAG sandwich ELISA
AUTHORS: Angel  Justiz-Vaillant

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This ELISA was used to study the interactions between Peptostreptococcal protein-L (SpL) and protein LAG (PLAG) with different immunoglobulin preparations from mammalian and avian species.</div></div>

[STEPS]
?. This ELISA was used to study the interactions between Peptostreptococcal protein-L (SpL) and protein LAG (PLAG) with different immunoglobulin preparations from mammalian and avian species. The 96 well microtiter plate was coated overnight at 4°C with 2 µg/µl per well of SpL in carbonate-bicarbonate buffer pH 9.6.
?. The plate was then treated with bovine serum albumin solution and washed 4X with PBS-Tween. 50 µl of immunoglobulins (1 mg/ml) is added and incubated for 1h at room temperature, and then, the microplate is rewashed 4X with PBS-Tween.
?. Then, 50 µL of peroxidase-labeled PLAG conjugate diluted 1:5000 in PBS-non-fat milk was added to each well and incubated for 1h at RT. The plate was washed 4X with PBS-Tween.
?. Then, 50 µL of o-phenylenediamine solution (4 mg/mL) was added, and the plate was incubated for 15 min at RT in the dark. The reaction was stopped with 50 µL of a 3M H2SO4 solution.
?. The plate was visually assessed for color development and read on a microplate reader at 492 nm. A cut-off point was calculated as the mean of the optical density of the negative controls multiplied by two. The cut-off point was set to 0.28.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
Identifying <em>putative</em> viral sequences from SPAdes-assembled data from the <a href="https://github.com/MicroB3-IS/osd-analysis" target="_blank">Ocean Sampling Day (2014)</a> metagenomic datasets using VirSorter.

[BEFORE_START]
To run this protocol, users must first <a href="https://user.iplantcollaborative.org/register/" target="_blank">register</a> for Cyverse account. All data (both inputs and outputs) are available within Cyverse's data store at /iplant/home/shared/iVirus/ExampleData/

[GUIDELINES]
This is part of a larger protocol <em>Collection</em> that involves the end-to-end processing of raw viral metagenomic reads obtained from a sequencing facility to assembly and analysis using Apps (i.e. tools) developed by iVirus and implemented within the Cyverse cyberinfrastructure.<br /><br />Finding putative viral sequences is one spring board for viral metagenomic analysis.

[STEPS]
?. 
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TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<div>
<div>
<div>
<p>During this step, you will be assembling the ARMS units from the individual parts. The unit is a stack of alternating open layers and closed layers. The closed layers contain a cross-shaped structure dividing the ARMS in four compartments, providing 'cave-like' spaces for organisms. The very first space, at the bottom of the ARMS <strong>should always be a closed layer</strong> and the top layer will be open.</p>
<p>Another important feature is that the first plate should lay flat on the base plate. No space should be created between the first plate and the base plate in order to avoid creating an extra space for colonization.</p>
</div>
</div>
</div>

[GUIDELINES]
<p>The numbering of the ARMS plates starts with the bottom plate, directly in contact with the base plate. This bottom plate is plate #1. The upper plate is plate #9.</p>
<p>The very first space, at the bottom of the ARMS <strong>should always be a closed layer</strong> and the top layer will always be open.</p>
<p>The first plate should lay flat on the base plate. No space should be created between the first plate and the base plate in order to avoid creating an extra space for colonization.</p>

[STEPS]
?. 
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TITLE:  Indirect Enzyme Linked Immunosorbent Assay (ELISA) for Detection of Anti-HIV Antibodies in Human Serum
AUTHORS: Angel  Justiz-Vaillant, Norma McFarlane-Anderson

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This protocol was already used successfully to detect anti-HIV antibody in the serum of women with cervical dysplasia or cervical cancer in Jamaica, West Indies [1].</div><div class = "text-block">Reference</div><div class = "text-block">1. Justiz Vaillant A, Bazuaye PE, McFarlane-Anderson N, Smikle MF et al. Seroprevalence of Anti-HIV Antibodies in Women with Abnormal Pap Smears in Jamaica. British Journal of Medicine & Medical Research 2013, 3(4): 2197-2202.</div></div>

[STEPS]
?. The 96 well polystyrene microplate (U-shaped bottom; Sigma-Aldrich) is coated with 50 ng of a mixture of synthetic peptides (including the fragment 579-601 of the HIV gp41 and fragments 254-274, 308-331 and 421-438 of the HIV gp120) for 4 h at 37ºC.
?. The microplate is blocked with 3% non-fat milk in PBS, 25 µl/well, 1h at room temperature (RT).
?. The microplate is washed 4X with PBS-Tween-20.
?. Duplicates of 25 µl of 1:16 diluted human sera are added.
?. After incubation for 90 min at RT the microplate is washed 4X with PBS-Tween 20.
?. Then, 25 µl of a chimeric commercially-prepared recombinant protein LA-HRP conjugate (Sigma-Aldrich) diluted 1:5000 is added.
?. After incubation for 90 min at RT and rewashing steps 25 µl TMB  is added to each well for 15 min in the dark.
?. The reaction is stopped with 3M H2SO4.
?. The microplate is read in a microplate reader at 450 nm.
?. In the ELISA is included a pooled human sera with high titre of anti-HIV antibodies as positive control, a pooled sera from healthy individuals as negative control and 0.9% normal saline solution was used as the blank.
?. The cut-off point is calculated as mean optical density (XOD) of negative control plus two standard deviation (SD).
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
Version February 14th, 2012

[STEPS]
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TITLE: JAX-Sen: Mice selection criteria
AUTHORS: Ramalakshmi Ramasamy, Juliana Alcoforado Diniz, Jessica Garofalo, Patrick Fleming, Paul Robson

[DESCRIPTION]
We aim to select 12 mice (C57BL/6), 6 male and 6 female at 6 and 12 months of age, 3 mice in each age and sex group. 

[STEPS]
TITLE: RT-QuIC alpha-synuclein 
AUTHORS: maria.xylaki

[DESCRIPTION]
This protocol is for the detection of prionoid alpha-synuclein forms in human cerebrospinal fluid using the Real Time Quacking-Induced Conversion method (RT-QuIC). The protocol is adapted from Marco J. Russo, Christina D. Orru, Luis Concha‑Marambio, Simone Giaisi et al., 2021 (doi:10.1186/s40478-021-01282-8) and Concha-Marambio et al., 2019 (doi:10.1007/978-1-4939-9124-2_4). 
This assay is for research use only and not diagnostics.

[STEPS]
SECTION: Preparation of Reaction buffer
2. Reaction mixture is prepared calculating 200 μl per well with the following final concentrations:
0.3 mg/mL recombinant alpha-synuclein
0.5 M NaCl
100 mM PIPES buffer
5 μM ThT
SECTION: Bead preparation
1. Beads are blocked with 1% BSA in 100mM PIPES for 1 hr and washed twice with PIPES.
SECTION: Seeting up the assay
3. One bead is placed in each well of the assay plate. 160 μl reaction buffer and 40 μl CSF are carefully pipetted in each well. Samples are assessed in triplicates.
SECTION: Seeting up the assay
4. Plate is covered with the film and creases are removed manually.
SECTION: Seeting up the assay
5. Plate is placed in the plate reader and incubated for 240 hrs at 37 °C in cycles of 1 min shaking at 500rpm, 29 mins incubation and fluorescence measurements are taken after every incubation cycle at 440ex/490emm.
SECTION: Data analysis
6. A sample is considered positive when it crosses a fluorescence threshold established at 3 standard deviations above baseline. 3/3 wells are positive and negative when 1/3 samples are positive while 2/3 is considered as incoclusive. 
SECTION: Data analysis
7. Relative fluorescence units measured at 490 are plotted versus the time in hours and should present a classical exponantial curve with lag phase and plateau. Kinetics parameters obtained from the curves include maximum fluorescence, time to reach 50% of maximal fluoresence and time to reach the established threshold.