eamag/protocols-with-steps huggingface dataset:
TITLE: Microfluidics Lithography 1 Mold Fabrication: Spin Coating of Photoresist (version s2)
AUTHORS:  C. Yunus Sahan, Serhat S

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><div class = "justify" style = "text-align:center"></div></div><div style = "text-align :; float : ;"><img style = "" src = "https://s3.amazonaws.com/pr-journal/hnnfjtn.png" /></div><br/><div class = "text-block"><div class = "justify" style = "text-align:center"><span>Nehir Biyoteknoloji Ltd. </span><a href="http://www.nehirbt.com.tr/en/" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">www.nehirbt.com</span></a></div></div></div>

[STEPS]
?. [SpinCoater Instrument Adjustment]
Dispense 1ml of photoresist (SU8) resin for each inch (25mm) of Si wafer substrate diameter.Spin at 500 rpm for 10 seconds with an acceleration of 100 rpm/second.Spin at "N" rpm for 30 seconds with an acceleration of 300 rpm/second.Expected results of photoresist thickness:   40um film thickness with N=4000 rpm spin of SU8 2050
   25um film thickness with N=4000 rpm spin of SU8 2025
   75um film thickness with N=2000 rpm spin of SU8 2050
   40um film thickness with N=2000 rpm spin of SU8 2025
The step is performed inside the Class 100.000 cleanroom and in a fume hood.
?. [Spin Coater Run]
Place the wafer in the spin coater and run the device.Microscope slides, Si/SiO2 wafers, and glass/PMMA wafers are alternatives substrates.
?. [Soft Baking of SU8 Coated Wafer]
Soft baking is done depending on the thickness of the coated photoresist film.It is a few minutes of baking at 65oC and 5-15 minutes of baking at 95oC are applied.
?. [Stocking the Coated Wafer]
In a petri plate, covered tightly with aluminum foil, photoresist-coated wafers can be stored in the cleanroom at room temperature for approximately one month.
TITLE: Visium Direct Mount Fresh Frozen -- University of Minnesota TMCs
AUTHORS: Laura J Niedernhofer, David A Bernlohr

[DESCRIPTION]
The Visium Spatial Gene Expression Solution measures the total mRNA in tissue sections and requires a Visium Spatial slide with intact tissue sections as input. This protocol outlines methanol fixation, Hematoxylin & Eosin (H&E) staining, and imaging of tissue for use with 10x Genomics Visium Spatial protocols. Fixed and stained tissue sections are inputs for the downstream Visium Spatial Tissue Optimization and Visium Spatial Gene Expression workflows.

Tissue sections are processed with Visium Spatial Gene Expression assay performed by the University of Minnesota Imaging Center and sequenced by the University of Minnesota Genomics Center.

[STEPS]
SECTION: Tissue Optimization
1.   

SECTION: Fixation, H&amp;E Staining &amp; Imaging
2.   

SECTION: Library Preparation &amp; Sequencing
3.   

 

SECTION: FASTQ Generation
4. BCL data from Illumina sequencer is demultiplexed and converted into FASTQ format using bcl2fastq version 2.20.0
TITLE: m9_media
AUTHORS: Elizabeth Fozo

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">M9 media protocols</div></div>

[STEPS]
?. [M9 minimal media: low osmolarity media for E. coli, resulting in slower growth rate]
Mix the ingredients (forms white precipitate but is dissolved after steering), filter sterilize, and store at room temperature.M9 Salts1X5X200100Magnesium sulfate2mM1M21Calcium chloride0.1mM1M0.10.05Vitamin B11ug/mL10mg.mL0.10.05Carbon sourceas requiredas requiredDistilled waterup to 1000mLup to 500mL
M9 Salts1X5X200100Magnesium sulfate2mM1M21Calcium chloride0.1mM1M0.10.05Vitamin B11ug/mL10mg.mL0.10.05Carbon sourceas requiredas requiredDistilled waterup to 1000mLup to 500mL
?. [Cold Spring Harbor Protocols:]
http://cshprotocols.cshlp.org/content/2006/1/pdb.rec8146.full?text_only=true
TITLE: Sample Informed Consent Form Template (Appendix C of Phase 3 study of Vaccine Candidate for COVID-19)
AUTHORS: Chris Ockenhouse, Chris Gast, Renee Holt, Jorge Flores

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span>This is Appendix C of "Phase 3 randomized, double-blinded, placebo-controlled trial to evaluate the safety, immunogenicity, and efficacy of </span><span style = "font-weight:bold;">Vaccine Candidate</span><span> against COVID-19 in adults > 18 years of age"</span></div><div class = "text-block">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.</div><div class = "text-block"><span style = "font-weight:bold;">Please note</span><span> that this is an evolving document, to be versioned and updated, based on community feedback and new data.</span></div></div>

[STEPS]
TITLE: Login to UA HPC web-portal
AUTHORS: Bonnie Hurwitz

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This describes the steps involved in logging into the UA High-Performance Computing (HPC) web-portal for the UA Biosystems Analytics class. The instructors should have already given you access to the HPC, if not contact us.</div></div>

[STEPS]
?. Go to the HPC web-portal: https://ood.hpc.arizona.edu/pun/sys/dashboard and login with your UA net-id
?. On the top menu bar select "clusters" and "ocelote shell access" from the pull-down list.
?. A shell terminal will open for you. You can navigate to the class directory using the "cd" command, and then create your own directory using the "mkdir" command (using YOUR net-id as the name):  $ cd /rsgrps/bh_class  $ mkdir my-netid
TITLE: The Simulated Random Assignment of Missense Mutations Throughout a Gene of Interest Can Determine Whether Missense Mutations Found in That Gene in a Population of Tumor Genomes Are Non-Randomly Distributed
AUTHORS: Richard L Cullum, David J Riese II

[DESCRIPTION]
Human malignancies result from the accumulation of genetic and epigenetic changes to normal cells. In many malignancies, gain-of-function mutations in oncogenes and loss-of-function mutations in tumor suppressor genes drive tumorigenesis and tumor progression. The identification of tumor driver mutations and the genes that host such mutations is critical for the molecular staging and targeted therapy of malignancies.

Since tumor driver mutations cause tumorigenesis or tumor progression, the proliferation of tumor cells selects for these mutations. Thus, in a gene that hosts tumor driver mutations, there will be a non-random distribution of mutations across the gene, as mutations that provide a selective advantage for the tumor cells will predominate over mutations that do not provide a selective advantage for the tumor cells.  Consider a particular gene in a population of tumor genomes; the total number of coincident missense mutations in that gene, defined here as two or more missense mutations that affect a particular codon, will be greater than the total number of coincident missense mutations that arise through random assignment of missense mutations across the gene.

Consequently, here we use the R Statistical Computing environment to simulate the random assignment of missense mutations across a user-specified gene.  The number of randomly assigned missense mutations is defined by the user and should be equal to the total number of missense mutations observed in the desired gene in the collection of tumor genomes of interest.  Based on the simulated random assignment of missense mutations, the R code then determines the total number of simulated coincident and non-coincident mutations.  This simulation is repeated a user-defined number of times, and the average number of simulated coincident and non-coincident mutations is calculated from the set of simulations.  

The R code then uses a Chi-square test to determine whether the observed number of coincident mutations (in the gene of interest in a collection of tumor genomes) significantly exceeds the average number of simulated coincident mutations.  A positive result indicates that the gene hosts a non-random distribution of missense mutations and suggests that the gene hosts tumor driver mutations.

We have used this R code to analyze mutations in the ERBB4 receptor tyrosine kinase gene that are found in The Cancer Genome Atlas (TCGA) dataset.  Our analysis indicates that the number of coincident mutations observed in ERBB4 in the TCGA dataset is statistically greater than the number of coincident mutations that arise from the simulated random assignment of missense mutations across the ERBB4 gene.  This finding indicates that the distribution of missense mutations in ERBB4 in the TCGA dataset is non-random.

[STEPS]
TITLE: Human Pancreas Histopathology Assessment
AUTHORS: Martha Campbell Thompson

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span style = "font-style:italic;">DRAFT</span></div><div class = "text-block">Purpose:</div><div class = "text-block">The purpose of this protocol is to outline proper procedures for fixed specimen processing using an automatic paraffin processor and sectioning and staining for histopathological review of normalcy. </div><div class = "text-block">Scope:</div><div class = "text-block">This protocol applies to human pancreas specimens that will be processed to paraffin blocks for HuBMAP Tissue Mapping Centers.</div><div class = "text-block">Expected Outcome:</div><div class = "text-block">Pancreas samples are processed properly and efficiently to whole slide images for histopathology review. </div><div class = "text-block">Nuclei should stain blue. Cytoplasm of acinar cells should stain pink with blue zymogen granules. The large blood vessels should demonstrate clear differentiation of RBCs when present, surrounding collagen, and endothelial nuclei with scant cytoplasm.</div></div>

[STEPS]
?. Process human pancreas for fixed paraffin samples (dx.doi.org/xxx) and submit to the University of Florida Molecular Pathology Core (https://molecular.pathology.ufl.edu/) or another GLP-compliant histology laboratory for routine paraffin embedding.
?. From 1 block of each region, create 4 µm paraffin sections and apply to Superfrost Plus slide or suitable alternative.
?. Stain sections with H&E (dx.doi.org/xxx).
?. Scan slides with whole slide digital scanner at 20x and save in default file format (Aperio .svs or Zeiss .cvs) or other lossless format (.tiff).
?. Optional: analyze tissue sample using ImageScope or QuPath for total section area, % fat infiltration.
?. Assess and record the following qualitative information for each section:1. Endocrine pancreas:     a. Islet numbers (normal, reduced, increased), sizes (normal range from small to large but majority medium sized), and morphology (normal oval outline with clear demarcation from surrounding acinar and duct cells).         i. Yes/No: amyloid, fibrosis, hydropic degeneration.2. Exocrine pancreas:     a. (0=none, 1=mild, 2=moderate, 3=severe) pancreatitis (subtype: acute, chronic), autolysis, fibrosis, fatty infiltration, neoplasia (including PanIN)3. Additional features:    a. Vascular network: arteriosclerosis, calcification vessels    a. Excretory system: duct dilation, protein plugging, stones
?. Based on histopathological evaluations, pancreas will be used when:All islet and exocrine features are scored at 0-1No pancreatitis
?. Inventory and archive paraffin blocks and H&E slides at room temperature.
?. Quality control:  Processor and staining solutions are changed at regular intervals to ensure adequate processing and staining of tissues. Temperatures and solution changes/rotations are recorded on the Paraffin Processor Use and Maintenance form. .
TITLE: Preparing DNA samples from mixed sand prior to extraction
AUTHORS: 

[DESCRIPTION]
<div class = "text-blocks"></div>

[STEPS]
?. Add 20 mL MilliQ water to pot
?. Remove plant, if any. Transfer to square petri dish.
?. Push a 50mL Falcon into the sand up to the 35 mL mark.
?. Tilt pot to a horizontal position and remove the falcon tube, retaining as much sand as possible.
Rotating the tube while removing it may help
?. Cap and wipe down the tube exterior using a paper towel with 70% EtOH before labelling with pot ID.
?. Fill tube with sand to the 40mL mark with MilliQ water
?. Vortex for 30 seconds
?. Pour through Miracloth into new 50ml tube. Discard the old tube.
?. Centrifuge tube at 5000 x G for 5 minute.
?. Pour away supernatant.
?. Add 1ml water and vortex to resuspend.
?. Transfer to 1.5ml tube for extraction or storage. Dispose of the 50ml tube.
?. Top up tube to 40 ml total volume with ultrapure water.
TITLE: Bogus Data Acquisition Protocol IV
AUTHORS: Abby   Moore

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

[BEFORE_START]
Before you start .....

[GUIDELINES]
Guidelines ....        

[STEPS]
1. In this step, I'll refer to the software that you need to use. I used the Software component to do this.


 

2. In this step, I'll refer to an image using embed code from Box.


 

3. In this step, I need to direct to to execute one of the following protocols depending on your context. Then you need to return to this protocol and continue with the next step. The two protocol options are hyperlinked below.


Bogus data acquisition protocol II
Bogus Data Acquisition Protocol III

4. Run the command displayed below, which is display with the command component.

 


5. In 
 
run the script display below with embed code.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
These methods accompany the following publication:<br /><br /><em><span style="color: #222222; font-family: arial, sans-serif; font-size: 12.8px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: normal; orphans: auto; text-align: start; text-indent: 0px; text-transform: none; white-space: normal; widows: 1; word-spacing: 0px; -webkit-text-stroke-width: 0px; display: inline !important; float: none; background-color: #ffffff;">Hebert, F, O; Grambauer, S; Barber, I; Landry, C, R; Aubin-Horth, N (2016): Reference transcriptome sequence resource for the study of the Cestode Schistocephalus solidus, a threespine stickleback parasite. GigaScience Database. </span><a href="http://dx.doi.org/10.5524/100197" target="_blank">http://dx.doi.org/10.5524/<wbr />100197</a></em>

[STEPS]
?.
TITLE: SDS-PAGE gel electrophoresis
AUTHORS: Lynn   Doran, Steven J Burgess

[DESCRIPTION]
SDS-PAGE gel electrophoresis protocol for analyzing samples from plant leaf tissue via immunofluorescence. In this protocol no Coomassie blue is added to samples, the reason is that this interferes with the fluorescent signal during immunoblot. Instead, samples have already been prepared in Laemmli buffer (minus coomassie, protein extraction procedure), the leading edge of samples can be visualized due to the presence of chlorophyll.  

Note 
- When using 15 well, 0.75 mm comb, try to limit the volume loaded to 10 μL to minimize the risk of spillover of protein between wells. 
- Ensure that accurate volume is pipetted by removing sample stuck to the outside of the pipette tip by wiping the tip on the rim of the sample tube to remove any residual liquid.


Literature: 
http://www.bio-rad.com/webroot/web/pdf/lsr/literature/Bulletin_6040.pdfhttps://www.bio-rad.com/webroot/web/pdf/lsr/literature/10026447.pdf

[BEFORE_START]
Extract protein from samples via Leaf Protein Extraction for Immunoblot  and determine Protein Concentration using Qubit 4 Fluorometer.

Previously extracted and quantified protein samples can be stored at -20 °C.

[STEPS]
SECTION: Prepare gel tank and buffers
1. Create a 1X working dilution of  Tris/Glycine/SDS buffer (~1 L  is required per gel tank) by diluting 10X stock 1:10 with distilled H2O.

SECTION: Prepare gel tank and buffers
2. Carefully remove the comb from the precast gel and the tape across the bottom. 



 


SECTION: Prepare gel tank and buffers
3. Assemble the Mini-PROTEAN electrophoresis cell and fill the inner chamber with buffer and the outer chamber up to the recommended mark 
 

SECTION: Prepare gel tank and buffers
4. Wash the wells with running buffer by pipetting up and down
 

SECTION: Prepare Samples
5. In fresh microcentrifuge tubes, create a dilution of each sample using 1x PEB to a pre-determined concentration total soluble protein. 
 


 

SECTION: Prepare Samples
7. Load 3 µL of Chameleon™ Duo Pre-stained Protein Ladder to the first well.

SECTION: Prepare Samples
8. Load 10 µL of each sample (30 µg of total soluble protein) per lane.



 



SECTION: Running Gel
9. Run precast gels at 170V for ~50 min or until the samples have reached the end of the gel. In some applications it may be advantageous to run the chlorophyll off the end of the gel to improve fluorescence and signal on the protein of interest.

 

 

SECTION: Running Gel
10. Carefully open precast gel case using an opening lever, by inserting where the black arrows indicate on the gel case.  Use the green Bio-rad scraper to cut the wells off of the gel.  Also use the scraper to remove the bottom edge of the gel that was below the union of the two pieces of plastic.
SECTION: Running Gel
11. Proceed either directly to Protein Transfer using Bio-rad TransBlot Turbo  or  Total Protein Staining.

SECTION: Prepare Samples
6. Briefly vortex the sample to shear any DNA contamination.
TITLE: In vitro transcription of DIG-labelled RNA probe
AUTHORS: Ida Barlow

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span>Protocol for making DIG-labelled RNA probes suitable for </span><span style = "font-style:italic;">in situ</span><span> hybridisation</span></div></div>

[STEPS]
?. Assemble reaction on ice:make up to 20ul total volume with WATER
[DTT]
[10X DIG-NTP mix (5 mM)]
[RNAse inhibitor]
[10 X transcription buffer]
[linearlised template]
[RNA polymerase]
[TOTAL volume]
5mM DIG-NTP mix (10X):
?. Mix well by gently flicking and spin down tube contents
?. Incubate at  for
37 °C
?. Add  and digest the template at  for
[DNaseI]
37 °C
?. Use ZYMO RNA cleanup kit to purify RNA and check quality by agarose gel electrophoresis; quantiy with nanodrop spectrophotometer
TITLE: RAPID RNA Extraction
AUTHORS: Noah Toppings

[STEPS]
?. Add 140 µL saliva to 560 µL lysis buffer.
?. Shake the tubes violently by hand.
?. Incubate at 61 ºC for 5 min.
?. Apply entire 700 µL to spin column.
?. Spin at 8 kRCF for 1 min.
?. Remove flowthrough with a pipette.
?. Add 500 µL wash 1 to the column.
?. Spin at 8 kRCF for 1 min.
?. Remove flowthrough with a pipette.
?. Add 500 µL wash 2.
?. Spin at 8 kRCF for 2 min.
?. Transfer column to a new tube and add 50 µL elution buffer.
?. Spin at 8k RCF for 1 min.
?. Add to LAMP reactions.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>This protocol refers to a study having the aim to identify and test at field artificial structures suitable as collectors for common cuttlefish <em>Sepia officinalis</em> eggs in wild condition.</p>
<p>During the same study, three different protocols were tested and compared. This one refers to the named 'trap' collector protocol, due to the adoption of trap structure used for fisheries.</p>
<p>This work was supported by the project “Protection, improvement and integrated management of the sea environment and of cross-border natural resources – ECOSEA”, funded by EU Adriatic IPA Cross-border Cooperation 2007-2013 (2°ord./0236/0).</p>
<p> </p>

[BEFORE_START]
<p>An adequate vessel for transshipment/positioning and scuba divers are needed for a proper installation.</p>

[GUIDELINES]
<p>The trap eggs collector consisted of a galvanized-iron frame (dimensions 80 x 50 x 30 cm) containing a 10 m long polyethylene floating rope (diameter 0.8 cm) wrapped around and acting as a substrate for cuttlefish deposition. This structure was similar to the traps commonly used by fishers, with the exception that this one was not hanged with a net so allowing the escapement of cuttlefish. A total of 111 collectors were deployed along the Marche region (Italy) in March, at the beginning of the spawning season, and kept at sea until July. During this period the collectors are regularly checked in order to evaluate the common cuttlefish eggs coverage.</p>
<p>This work was supported by the project “Protection, improvement and integrated management of the sea environment and of cross-border natural resources – ECOSEA”, funded by EU Adriatic IPA Cross-border Cooperation 2007-2013 (2°ord./0236/0).</p>
<p> </p>
<p> </p>
<p><img id="s-mce-img" class="s-mce-img" src="https://s3.amazonaws.com/pr-journal/5d82xfe.png" data-src="https://s3.amazonaws.com/pr-journal/5d72xfe.png" data-ofn="Trap collector.png" /></p>
<p> </p>
<p> </p>
<p><img id="s-mce-img" class="s-mce-img" src="https://s3.amazonaws.com/pr-journal/5ea2xfe.jpg" data-src="https://s3.amazonaws.com/pr-journal/5d92xfe.jpg" data-ofn="Trap collector.jpg" /></p>

[STEPS]
?.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
Importance
Recently, we showed that the elderly are significantly underrepresented in randomized controlled trials (RCT) in rheumatoid arthritis (RA) and osteoarthritis (OA). While this phenomenon has been detected in other fields as well, efforts by various international institutions to tackle the issue were without decisive success. Researchers might be cautious about including more elderly people because they fear reduced trial retention rates.
Objective
To evaluate whether the proportion of included elderly individuals (defined by an age of ≥65 years) is independently associated with trials’ retention rates.
Data Source
MEDLINE (via PubMed).
Eligibility
RCT on any intervention in RA or OA published in 2016 or 2017.
Outcome
Retention rate.
Critical Appraisal
We will not address any conclusion made by included RCT. Thus, we will not perform a formal risk of bias assessment.
Synthesis Methods
The proportion of elderly people is either directly abstracted from the research manuscript or estimated from an assumed truncated normal distribution. Multivariable meta-regression will explore whether the proportion of included elderly people is independently associated with trials’ retention rates, even after adjusting for trial duration. The model will include as covariates – apart from the proportion of elderly people and study duration - disease, type of intervention, region, sample size at enrollment, and the proportion of women. Additional models will explore whether the proportion of included elderly people is independently associated with trial retention rates when only drop-outs due to adverse events, resp. lack of efficacy are counted.

[GUIDELINES]
PRISMA
Preferred Reporting Items for Systematic Reviews and Meta-Analyses

Liberati A, Altman DG, Tetzlaff J, et al. The PRISMA statement for reporting systematic reviews and meta-analyses of studies that evaluate health care interventions: explanation and elaboration. Ann Intern Med 2009;151(4):W65-94.

PRISMA-P
Preferred reporting items for systematic review and meta-analysis protocols

Shamseer L, Moher D, Clarke M, et al. Preferred reporting items for systematic review and meta-analysis protocols (PRISMA-P) 2015: elaboration and explanation. BMJ 2015;349:g7647. doi: 10.1136/bmj.g7647.

[STEPS]
SECTION: INTRODUCTION
?. 
SECTION: RATIONALE
?. 
SECTION: METHODS
?. 
SECTION: ETHICS AND DISSEMINATION
?. 
SECTION: COMPETING INTERESTS
?. 
SECTION: FUNDING
?. 
SECTION: REFERENCES
?.