eamag/protocols-with-steps huggingface dataset:
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>this experiment describes a process to deveop a classifier which can be used to modulate the stiffness of prosthetic leg based on the movement of hands, elbows and position of the other leg of the amputee</p>

[GUIDELINES]
<p>the initial experiments shoud be conducted on healthy subject.</p>

[STEPS]
?. 
?. 
?. 
?. 
?. 
?.
TITLE: My shiny new protocol
AUTHORS: Emma  Ganley

[DESCRIPTION]
test abstract

[STEPS]
SECTION: Section 1
1. 

step 1
SECTION: Section 1
2. add 50 mL and 100 mL

SECTION: section 2
4. 2 min 

tesating


SECTION: Section 1
3. 
SECTION: section 2
5. a new step
SECTION: Section 3
6.
TITLE: FibPho: Analysis Protocol
AUTHORS: Sasha Burwell

[DESCRIPTION]
This protocol details the analysis of the fiber photometry data.


[STEPS]
SECTION: Analysis Protocol
1. Each animal has one folder per recording session generated by Synapse with all the fiber photometry recording data from that session. Make a copy of the 
 
 generated by the behavior code from that same session and add it to the folder.
SECTION: Analysis Protocol
2. To analyze the fiber photometry signals time-locked to the behavioral trials, across multiple behavior sessions for one animal, run the MATLAB code 
 

This saves a 
 
 variable with the raw 415nm and 465nm signals and the calculated dF/F signal per behavior trial
SECTION: Analysis Protocol
3. Copy the 
 
 variables from all mice in an experimental condition into one folder
SECTION: Analysis Protocol
4. Run the MATLAB code 
 
 to combine and analyze trial-based fiber photometry signals across animals.
SECTION: Analysis Protocol
4.1. Modify the 
 
variable (line 39) to generate the data for the specific session/trials you are interested in.
SECTION: Analysis Protocol
4.2. This code calculates a mean dF/F signal per animal for the cue and reward time windows, and prints them to the command line.
SECTION: Analysis Protocol
4.3. This code also generates the across time waveform plots for visualization of mean and SEM, as in Figure 3 and Ext. Figure 4.
SECTION: Analysis Protocol
5. Use the MATLAB code 
 
 to analyze the difference between groups and generate Ext. Fig 4b.
SECTION: Analysis Protocol
5.1. X values = cue-burst sessions 6-10 (
 
).
SECTION: Analysis Protocol
5.2. Y values = reward-omission pause first 4 extinction trials (
 
).
SECTION: Analysis Protocol
5.3. Exclude mice with recording signals too low to detect pauses (X < 1).
SECTION: Analysis Protocol
6. Use the MATLAB code 
 
 with the mean dF/F values of the included mice (calculated above) to generate the Gardner-Altman plots in Figure 3 and Ext. Figure 4
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>Infected beetles were collected and homogenized in sterile water. The homogenates were filtered through four layers of cheesecloth and centrifuged at 3000g for 15 min. The pellets were resuspended in sterile water, and the spores were purified by Percoll gradient centrifugation using 90% Percoll at 15,000g for 40 min. The spore band was collected and washed several times with sterile water. The purified spores (n = 50) were measured under a light microscope (IL/Leica Microsystems, Inc., Deerfield) with an ocular micrometer and photographed with the Microscope USB Camera.</p>

[STEPS]
TITLE: Registro y entrada de material bibliográfico Biblioteca de la UCAM
AUTHORS: Antonio Rex Alegria

[DESCRIPTION]
Registro y entrada de material bibliográfico Biblioteca de la UCAM

[STEPS]
1. Abrimos nuestra página de Excel donde introducimos los datos correspondientes al ejemplar que registramos en la Biblioteca

 
     Nº registro      Fecha Reg.      Autor      Título      C.D.U.             
 
2. En la portada del material, ponemos el sello de la Biblioteca y escribimos el número correspondiente de la hoja de Excel.

  

3. Una vez registrado, catalogamos el ejemplar con el programa de gestión Bibliotecaria Absysnet.


  

4. Una vez catalogado el material, colocamos el sistema anti-hurto y el tejuelo en el lomo del ejemplar.

  

5. Colocamos el código de barras, encima del sello del registro.    

6. Por último, colocamos el ejemplar en la estantería.
TITLE: Peptide fragment (579-601 from HIV-gp41) conjugated to keyhole limpet haemocyanin to be used as HIV immunogen.
AUTHORS: Angel  Justiz-Vaillant

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">Chemical synthesis facilitates the generation of peptides which are exceedingly difficult to express in bacteria, peptide/protein backbone modification, the incorporation of unnatural amino acids, and the production or synthesis of D-proteins. </div><div class = "text-block">The C-terminal cysteine can be added to the amino acidic sequences of HIV peptides (fragment 579-601 of the HIV-gp41 [1]. These peptide fragments were dimerized by cysteine oxidation with dimethyl-sulfoxide [2] to facilitate their conjugation to keyhole limpet hemocyanin that acts as a carrier protein. </div><div class = "text-block">Reference:</div><div class = "text-block"><ol style = "list-style-type: decimal;"><li style = "counter-reset:ol0;"><span>McPhee DA, Kemp BE, Cumming S, Stapleton D, Gust ID, Doherty RR. Recognition of envelope and tat protein synthetic peptide analogs by HIV positive sera or plasma. </span><span style = "font-style:italic;">FEBS Lett</span><span>. 1988;233(2):393-396. doi:10.1016/0014-5793(88)80468-x. </span></li><li style = "counter-reset:ol0;">Tam JP, Wu CR, Liu w, Zhang JW (1991) Disulfide bond formation in peptides by dimethyl sulfoxide. Scope and applications. J Am Chem Soc 113: 66576662. </li></ol></div></div>

[STEPS]
?. These peptide fragment (579-601 from HIV-gp41) is dimerized by cysteine oxidation with dimethyl-sulfoxide. The HIV peptide is dissolved in 5% acetic acid to a final concentration of 5.1 mg/ml.
?. The pH of the medium is adjusted to 6 with 1 M (NH4)2CO3.
?. Dimethyl-sulfoxide is added to 20% of the final volume, and after four hours at room temperature (RT), the solute is extracted.
?. Then, the peptide is dissolved in 3 ml 5% trifluoroacetic acid and precipitated with 35 ml cold ether.
?. The precipitate is dialyzed against 1.2 liters of deionized water, pH 7 at 4°C overnight.
?. The fragment 579-601 of gp 120 was conjugated by the glutaraldehyde method.
?. One mg of keyhole limpet hemocyanin (KLH) is dissolved in 2 ml 0.1 M borate buffer (1.24 g boric acid, 1.90 g sodium tetraborate, pH 10, in 500 mL deionized water).
?. In a 20 ml glass tube by gentle stirring 1 µmol of the HIV synthetic peptide and 0.2 mL 0.3% glutaraldehyde solution (ACS reagent grade, pH 5.5, Sigma-Aldrich) are slowly mixed at RT and left to stand for 2 hrs.
?. When a yellow coloration is observed this indicates that the conjugation process is successful.
?. To blocking the excess of glutaraldehyde, 0.26 ml of 1 M glycine (Sigma-Aldrich) is added.
?. The mix is left for 32 min at RT.
?. The HIV-hemocynin conjugate is then dialyzed against 1.3 liters 0.1 M of borate buffer, pH 8.4 through the night at 4°C.
?. Then the 0.1 M borate buffer is used to dialyze the preparations for 8 hrs at 4°C.
TITLE: PNA-functional Ligand Conjugates Synthesis
AUTHORS: Cathy  Miller

[DESCRIPTION]
In recent years, the development of PNA into genetic drugs has been one of the research hotspots. However, the biggest obstacle leading to the poor drug ability of PNA is still the problem of cell transport. Covalently linking PNA with a suitable functional ligand with pharmacological activity is a successful attempt in the development of PNA drugs. The successful design of PNA-functional ligand conjugates not only improves the cellular uptake of PNA, but also introduces functional ligand compounds into cells to achieve dual therapeutic effects of PNA and functional ligands. In this conjugate, the ligand not only has the function of assisting the transport, but also exerts its own pharmacological activity.

At Creative Peptides, we always pay attention to the needs of our customers and recognize that careful planning and perfect execution are the key factors for success. Therefore, with our experience and our expertise in the PNA field, for each problem, we will form a professional development team to help you choose the process and technology that meets your research goals.

[STEPS]
TITLE: Incubation chamber settings
AUTHORS: Wolfram Moebius

[DESCRIPTION]
Incubation chamber settings

[STEPS]
SECTION: Chamber Settings For 30°C
1. 
Newer chamber:
Lid = 32 °C
Plate = 29 °C
Glass = 28 °C
Probe reading 30 °Cthroughout
Thermal camera image 2777


Older chamber:
Lid = 33.5 °C
Plate = 31 °C
Glass = 30 °C  
Probe reading 30 °Chroughout
Thermal camera image 2786

New chamber at 30 °C
 
Old chamber at 30 °C
 



SECTION: Chamber Settings For 37°C
1. Final Chamber values used afte 14/09/18:

Newer chamber:
Lid = 38 °C
Plate = 36 °C
Glass =35 °C
Probe reading36.8 °Cthroughout
Thermal camera image 2795


Older chamber:
Lid = 40.5 °C
Plate = 38 °C
Glass =37 °C
Probe reading 36.7 °Cthroughout
Thermal camera image 2790

New chamber at 37 °C:
 
 Old chamber at 37 °C:

 

smb://mcrlsinas01.ex.ac.uk/lab-moebius/cmaberly/Incubation Chamber Camera Images


SECTION: Chamber Settings For 37°C (OLD)
2. 
Newer chamber:
Lid = 38 °C
Plate = 36 °C
Glass = 35 °C
Probe reading 36.8 °Cthroughout
Thermal camera image 2795


Older chamber:
Lid = 40.5 °C
Plate =38 °C
Glass = 37 °C
Probe reading 36.7 °Cthroughout
Thermal camera image 2790

New chamber at37 °C:
 

Old chamber at37 °C:
 

smb://mcrlsinas01.ex.ac.uk/lab-moebius/cmaberly/Incubation Chamber Camera Images



SECTION: Chamber Settings For 37°C UPDATED VERSION
3. Newer chamber:
Lid = 38 °C
Plate = 36 °C
Glass = 35.5 °C
Probe reading 36.2 °Cthroughout

Older chamber:
Lid = 40.5 °C
Plate =38 °C
Glass = 37 °C
Probe reading 36.7 °Cthroughout
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
This collection is part of the VERVE holiday drive to collect exotic off-the-shelf laboratory recipes. More details <a href="https://www.protocols.io/g/verve-net/news/call-for-recipes" target="_blank">here</a>.<br /><br />If you know creative/unusual protocols like these, please tweet them to #ExoticMethods, e-mail to "<a href="mailto:info@protocols.io" target="_blank"><span class="s2">info@protocols.io</span></a>", or share directly as a protocol here, tagging with "ExoticMethods".

[STEPS]
?.
TITLE: Transformation Protocol for BL21(DE3) Competent Cells (C2527I)
AUTHORS: New England  Biolabs

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span>This transformation protocol is for the C2527I cells. (For the C2527H protocol, see </span><a href="https://www.protocols.io/view/Transformation-Protocol-C2527H-imsv4d" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">here</span></a><span>.)</span></div></div>

[STEPS]
?. Thaw a tube of BL21(DE3) Competent E. coli cells  until the last ice crystals disappear.
on ice
?. Mix gently and carefully pipette  into a transformation tube .
[cells]
on ice
?. Add – containing – to the cell mixture.
1 µl
5 µl
1 pg
[plasmid DNA]
?. Carefully flick the tube 4–5 times to mix cells and DNA. Do not vortex.
?. Place the mixture  for . Do not mix.
on ice
?. Heat shock at exactly  for exactly . Do not mix.
42 °C
?. Place  for . Do not mix.
on ice
?. Pipette  of   into the mixture.
950 µl
[SOC]
?. Place at  for . Shake vigorously () or rotate.
37 °C
Centrifuge: 250 33
?. Warm selection plates to .
37 °C
?. Mix the cells thoroughly by flicking the tube and inverting.
?. Perform several 10-fold serial dilutions in SOC.
?. Spread – of each dilution onto a selection plate and incubate  at .
50 µl
100 µl
37 °C
Alternatively, incubate at   for  –  or at   for  .
TITLE: Infection of Biomphalaria glabrata snails with  Schistosoma mansoni miracidia
AUTHORS: Sarah K Buddenborg

[DESCRIPTION]
To infect Biomphalaria glabrata snails with miracidia hatched from Schistosoma mansoni eggs

[STEPS]
SECTION: Lung preparation
1. Collect lungs from patent mice into 50ml Falcon tubes containing pre-warmed 37°C 1x DPBS
SECTION: Lung preparation
2. Remove lungs from PBS and place in large mortar or laboratory blender
SECTION: Lung preparation
3. Gently homogenise lung tissue
SECTION: Lung preparation
4. Place homogenised lung slurry into a flask (size of flask depends on how many lungs you have) and fill with diH2O or aquarium water
SECTION: Hatching miracidia
5. Cover flask in aluminum foil and shine light horizontally across the opening of the flask for 1-2 hrs
SECTION: Exposing snails to miracidia for monomiracidium infections
11. Dilute miracidia so that one miracidium can easily be collected in ~3-5µl of water
SECTION: Exposing snails to miracidia for regular life cycle maintenance
8. Place individual 5-8mm snails into 24-well culture plates and cover snails completely with 1x aquarium water
SECTION: Hatching miracidia
6. Take a small aliquot of water from the top of the flask using a glass Pasteur pipette and place into a petri dish. Using a stereomicroscope, check for swimming miracidia
SECTION: Exposing snails to miracidia for regular life cycle maintenance
10. After exposure, remove the snails carefully using featherweight forceps and put them in a new tank with food
SECTION: Exposing snails to miracidia for monomiracidium infections
12. Collected single miracidium using fresh 10µl pipette each collection and place in 24-well plate
SECTION: Exposing snails to miracidia for monomiracidium infections
13. After a plate is filled, check each well under a microscope to verify there is a single miracidium in each well
SECTION: Exposing snails to miracidia for regular life cycle maintenance
7. Under a microscope, collect 15 miracidia with a glass Pasteur pipette per well

Alternatively (but less preferred) estimate the number of miracidia by counting 12-5µl aliquots in a petri dish. After collecting (shaking the tube in between) and placing the aliquots in the petri dish add 5µl of Lugol to each drop (this kills and stains the miracidia) 
SECTION: Exposing snails to miracidia for monomiracidium infections
14. Place individual 5-8mm snails in the wells containing confirmed single miracidium and cover snails completely with 1x aquarium water
SECTION: Exposing snails to miracidia for monomiracidium infections
15. Leave the plate overnight (inside incubator or room at 28°C)
SECTION: Exposing snails to miracidia for monomiracidium infections
16. The following day, transfer the snails to a new tank with food
SECTION: Exposing snails to miracidia for regular life cycle maintenance
9. Expose snails to miracidia for at least 3 hours (up to overnight)
TITLE: Copy of Fluorescence analysis using CF imager-v2
AUTHORS: Steven Burgess

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

[STEPS]
?. The surface of the leaf should be 140mm from the base of the imaging chamber, and can be adjusted by lowering or raising the plant under analysis. Position plant/leaf in the chamber
?. [Set exposure]
Maually adjust the apeture as shown on the right to allow and optimal amount of light into the imager so as not to overexpose measurements.After adjusting the focus and press the "map image" button, change the plate/plant to the one you are going to read. If you don't press "map image", the light will be still flashing when you open the chamber.
?. Isolate the plant or leaf of interestApplying isolation background - it can be done more than once if the image is still with some backgroundAfter applying isolation background areas will be masked out in blue as shown below.
?. Remove residual noise from imageSometimes the software picks up background noise as real signals, shown below as black dots on the blue background. It is advisable to mask these, otherwise they will be counted as a separate 'colony' during analysis and measurements will be recorded for each of these spots in the final data sheet. Noise can be masked by moving the cursor over the dot, pressing CTRL+left click simultaneously.
?. Load protocol
?. If the protocol needs change, press Settings > Protocol. Use the left menu to adjust the parameters you want to.
?. 
?. Once you are happy with the scheduled program click on the protocol icon in the toolbar (black arrow below) to start the run
?. Export dataAfter the reading is complete, save the images (File > Save As). To obtain the files to process the images, go to File > Export to Folder. It will save in the folder that is opened.
?. [Preparing plants]
Dark adapt plants for at least 20 minutes prior to taking measurements.
This is done to ensure the photosynthetic electron transport chain is fully oxidized and reaction centres are open. In an ideal situation plants are allowed to dark adapt overnight prior to measurement.A properlly adapted, healthy plant should give a Fv/Fm value of ~0.8. This has been shown to be highly stable between species. Significant deviation from this (e.g. Prepare an extra plate with few leaf disks(or have extra plants/leaf disks) to adjust the focus (on step 5). During the focus adjustment, the light flashes inside the chamber, so you should not use the plates that are with yourdark-adapted samples.
?. Start the FluorImager software
?. Turn on the cf imager and open the FluorImager software.
?. [Set focus]
Before initiating this step, place the extra plate/plants inside the chamber.Set the focus by adjusting the dial above the chamber, and lock in position by turning the screw on the side.
TITLE: Passaging cells in MultiFlasks
AUTHORS: Allan JW Lui

[DESCRIPTION]
Simple protocol for working with Falcon Multi-Flasks

[STEPS]
SECTION: 5-layer Multi-Flask
1. Per flask, Prepare and pre-warm:
200ml PBS
30ml 0.25% Trypsin-EDTA or other detachment reagent
70ml complete media for trypsin inactivation, plus extra for reseeding
SECTION: 5-layer Multi-Flask
2. Pour out media in flask, wash out remaining media twice by:
Adding 100 mL
Distributing equally and evenly between layers to cover the entire culture surface area
Pouring out PBS


SECTION: 5-layer Multi-Flask
3. Add 30 mL and distribute equally between layers
Incubate 3 min 37 °C 

SECTION: 5-layer Multi-Flask
4. Inactivate trypsin with 30 mL distributed equally between layers
Collect cell suspension into 50ml tubes or 100ml bottles
Wash out remaining cells with 40 mL

SECTION: 5-layer Multi-Flask
5. Centrifuge at 200 x g, 5 min, remove trypsin, resuspend in fresh media if necessary, then count cells and reseed

6. Per flask, Prepare and pre-warm:
TITLE: Confocal microscopy and characterization of synaptic boutons associated with ganglion neurons
AUTHORS: Janet Keast, Peregrine Osborne

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This protocol describes confocal microscopy and image analysis procedures for characterizing neuronal cell bodies and their associated synaptic boutons in thick (50 µm) cryosections. The protocol has been applied to rat pelvic ganglia, where neuronal cell bodies have been identified using immunohistochemical markers of specific neuron populations and/or fluorescent retrograde tracer.</div></div>

[STEPS]
?. [Confocal microscopy]
Using Zen Black (Zeiss software), batch process Airyscan output to 16-bit 1260 x 1260 pixel images.
?. [Confocal microscopy]
To optimize visualization of synaptic boutons, image using a 60x 1.4 NA oil immersion objective with 3x digital zoom. Multi-channel images were collected by sequential imaging of channels, with four scans averaged per image.
If a particular channel is susceptible to photobleaching (e.g., retrograde tracers such as FluoroGold), image this channel first.For these studies, a Zeiss LSM880 was used in Fast Airyscan mode.
?. [Image analysis]
Airyscan processed images were converted to Imaris (.ims) files for analysis using the Surface Rendering module.
?. [Image analysis]
For the channel showing the neuronal cell body of interest, create a mask by first creating the neuronal surface and processing in a defined volume. Use parameters tailored to your specific immunolabeling signal requirements. Examples of parameters for manual adjustment include background subtraction, split touching objects and threshold.
?. Delete all other objects from this channel, select a surface of the neuron and merge all surfaces to create an appropriate mask of the neuron.
For studies of pelvic ganglion cell bodies, surface detail is set to 0.2 µm.
?. For the channel showing synaptic boutons, create mask and surface of the boutons as above.
For studies of synaptic boutons associated with pelvic ganglion, surface detail is set to 0.05 µm.
?. To analyse the distance between the neuronal cell body surface and the synaptic boutons, apply the Distance Transformation extension. This calculates the closest distance between two surfaces, and the surface area and volume of all objects with rendered surfaces.
?. Export data separately from each channel (neuronal cell body, boutons) to Excel.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<span class="cit-title">This protocols is from:<br /></span><span class="cit-auth cit-auth-type-author">Linlin Yin</span><span class="cit-sep cit-sep-separator">, et al.</span><span class="cit-auth cit-auth-type-author"> (2015) <a href="http://www.genetics.org/content/200/2/431.long" target="_blank">Multiplex Conditional Mutagenesis Using Transgenic Expression of Cas9 and sgRNAs</a>. </span><em>Genetics </em><span class="cit-vol">200<span class="cit-sep cit-sep-after-article-vol">:</span></span><span class="cit-pages"><span class="cit-first-page">431</span><span class="cit-sep">-</span><span class="cit-last-page">441</span></span><span class="cit-elocation">; </span><span class="cit-doi"><span class="cit-sep cit-sep-before-article-doi">doi:</span>10.1534/genetics.115.176917<br /><br />Please see the<a href="http://www.genetics.org/content/200/2/431.long" target="_blank"> full manuscript</a> for additional details.</span>

[GUIDELINES]
<img id="s-mce-img" class="s-mce-img" src="https://s3.amazonaws.com/pr-journal/4rcz36.png" alt="" data-src="https://s3.amazonaws.com/pr-journal/4qcz36.png" data-ofn="Screen Shot 2015-09-07 at 9.53.47 PM.png" /> <br /><br />-------------------<br />List of plasmids deposited at Addgene (Deposit 71794)<br /><br /><strong>Plasmid ID Plasmid Name</strong> <br />64237 pME-Cas9 <br />64239 pGGDestTol2LC-1sgRNA <br />64240 pGGDestTol2LC-2sgRNA <br />64241 pGGDestTol2LC-3sgRNA <br />64242 pGGDestTol2LC-4sgRNA <br />64243 pGGDestTol2LC-5sgRNA <br />64245 pU6a:sgRNA#1 <br />64246 pU6a:sgRNA#2 <br />64247 pU6b:sgRNA#3 <br />64248 pU6c:sgRNA#4 <br />64249 pU6d:sgRNA#5 <br />64250 pU6a:sgRNA(tyr)<br />------------------------<br /><strong>Reference</strong>:<br />JAO, L. E., S. R. WENTE and W. CHEN, 2013 <a href="http://www.pnas.org/content/110/34/13904.abstract" target="_blank">Efficient multiplex biallelic zebrafish genome editing using a CRISPR nuclease system</a>. Proc Natl Acad Sci U S A 110: 13904- 13909.<br /><br />

[STEPS]
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?.