eamag/protocols-with-steps huggingface dataset:
TITLE: Performance Study of Wireless Fecobionics Device in Canine
AUTHORS: Yanmin Wang, Hans Gregersen

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">We developed a novel wireless device (Fecobionics) for mapping colonic and anorectal neuromuscular function. The hypothesis of this protocol is that the Fecobionics device can test pressures, orientation, bending, shape, and cross-sectional area changes in colon and rectum. To validate our hypothesis, we inserted the device to the proximal colon, either through a cannula or colonoscopy, followed by data transmission and recording.</div></div>

[STEPS]
?. [Cannula test]
After a laparotomy, a cannula was implanted into the proximal of the colon (4-5 cm to cecum)
?. [Cannula test]
The external end of the cannula was screwed by a cap, which is able to open for device insertion
?. [Cannula test]
After 10-14 days recovery, the Fecobionics was inserted through the cannula for testing
?. [Cannula test]
X-ray fluoroscopy was performed frequently to locate the device. Data was recorded for analysis.
?. [Colonoscopy]
After 2-day laxative and an enema, the animal was anesthetize for colonoscopy
?. [Colonoscopy]
The Fecobionics was delivered to proximal colon (close to cecum) by an endoscopic rat-tooth forcep
?. [Colonoscopy]
After the delivery, X-ray fluoroscopy was performed frequently to locate the device. Data was recorded for analysis.
TITLE: No Title
AUTHORS: 

[GUIDELINES]
<p>Use with Ultra Streptavidin Detection Kit (<a href="https://antibody.biolegend.com/datasheet.php?UpProd=&amp;catalogno=SIG-32250" target="_blank">SIG-32250)</a> or (<a href="https://antibody.biolegend.com/datasheet.php?UpProd=&amp;catalogno=SIG-32248" target="_blank">SIG-32248)</a></p>
<p> </p>
<p>Positive control: Normal human cerebellum (except SMI-71, which should be rat brain)</p>

[STEPS]
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TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<section>
<p><a href="http://bioinformatics.psb.ugent.be/webtools/trapid/" target="_blank">TRAPID</a> is an online tool for the fast, reliable and user-friendly analysis of <em>de novo</em> transcriptomes, developed and maintained by the CNB group at VIB-UGent Center for Plant Systems Biology.</p>
<p> </p>
</section>
<p>This protocol explains how to perform functional annotation of a transcriptome using the TRAPID platform. It also describes basic analyses that can be performed within TRAPID, after initial processig of the data.</p>

[BEFORE_START]
<p>Make sure that:</p>
<ul>
<li>You are using a web browser with JavaScript and Adobe Flash enabled. Supported browsers: Safari, Chrome, Firefox (most recent versions).</li>
<li>Java is installed on your machine (version JRE 6.0 or higher). This is required to launch the JalView alignment editor and the Archeopteryx tree viewer.</li>
</ul>

[GUIDELINES]
<p>In this protocol, we work with an example dataset consisting in 25,392 transcript sequences from <em>Panicum hallii </em>(<a href="https://doi.org/10.1111/j.1365-313X.2012.04938.x" target="_blank">Meyer et al. 2012</a>). This dataset can be retrieved from TRAPID's FTP <a href="ftp://ftp.psb.ugent.be/pub/trapid/datasets/panicum" target="_blank">here</a>.</p>

[STEPS]
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TITLE: Classic Latke&#39;s Recipe
AUTHORS: Lenny Teytelman, Dmitry Lupyan

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span>This recipe is traditional Eastern-European Jewish latkes (potato pancakes). This latke recipe was adapted from </span><a href="https://cooking.nytimes.com/recipes/1015533-classic-potato-latkes" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">The New York Times</span></a><span>. It can be eaten with apple sauce or sour cream. </span></div></div>

[STEPS]
?. Do not peel!  No need to peel!  Wash 6 Russet potatoes.
?. With either food processor or hand grater, use the coarse disc/side to grate the potatoes and the onion into the bowl
?. Transfer potato/onion mixture into a cloth towel and squeeze all the moisture possible. (this is a critical step for the latkes to come out crispy).
?. Transfer the mixture back into the bawl
?. Add salt, pepper, baking powder and eggs, mix well.
?. Cover bottom of a frying pan with oil and heat to medium-high (about ¼ inch thick).
?. One tablespoon will yield one latke. Depending on the size of the pan, you'll be able to fit 6-10 latkes.Scoop up the mixture, place it on the hot pan. Turn the latke after 4-5 minutes (or when the edges turn golden brown). Fry for additional 4-5 minutes on the other side.
?. Place the latkes on the large plate lined with a paper towel. Sprinkle a bit of salt after every batch.Enjoy!
TITLE: Feacalis_colony_PCR
AUTHORS: Elizabeth Fozo

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span style = "font-style:italic;">E. faecalis</span><span> colony PCR using taq polymerase</span></div></div>

[STEPS]
?. [E. faecalis colony PCR using taq polymerase]
Transfer 1 colony, depending on size to 50-100µLwater. Normal size colonies can go in ~65µLwater.
?. [E. faecalis colony PCR using taq polymerase]
To a PCR tube add:15 µL colony water*1.25 µL primer 1 (20 µM stock)1.25 µL primer 2 (20 µM stock)2.5 µL10X thermopolbuffer0.5 µL10 mM dNTPs0.5 µL Taq polymerase4 µL waterTotal  = 25 µL
*Master mix can be made with all ingredients except the colony water for multiple reactions using the same primers.
?. [E. faecalis colony PCR using taq polymerase]
Use the following cycle in the thermocycler:94°C        10 min       1 cycle94°C         15 sec  54°C         30sec(5°C below Tm)72°C         1 min/kb     25-30 cycles72°C         10 min        1 cycle15°C          HOLD
TITLE: sci-RNA-seq2 Pipeline for the Human Biomolecular Atlas Program (HuBMAP)
AUTHORS: Shin Lin, Cole Trapnell, Jay Shendure

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This document provides an overview of the protocols used by the Caltech-UW TMC to perform sci-RNA-seq2. This involves sequencing of the transcriptomes of dissociated single cells in a massively parallel manner.</div></div>

[STEPS]
?. Collection and processing of human tissues.Sample procurement: dx.doi.org/10.17504/protocols.io.biqpkdvnFlash freezing: dx.doi.org/10.17504/protocols.io.biqqkdvw
?. sci-RNA-seq2 assaydx.doi.org/10.17504/protocols.io.binskdee
TITLE: Purification of mCherry-WIPI2d/WIPI3
AUTHORS: Elias Adriaenssens, Dorotea Fracchiolla

[DESCRIPTION]
This protocol details the purification of mCherry- WIPI2d/WIPI3.

[STEPS]
SECTION: Purification procedure
1. To purify mCherry-WIPI2d and mCherry-WIPI3, as described previously for WIPI2d (Fracchiolla et al. 2020, J Cell Biol, PMID: 32437499), fuse the coding sequence of WIPI2d or WIPI3 to a N-terminal 6xHis-TEV-mCherry-tag through cloning into a pET-DUET1 vector (available from Addgene). 

SECTION: Purification procedure
2. After the transformation of the pET-DUET1 vector encoding 6xHis-TEV-mCherry-WIPI2d/WIPI3 in E. coli Rosetta pLysS cells (Novagen Cat# 70956-4), grow the cells in 2x Tryptone Yeast extract (TY) medium at 37 °C until an OD600 of 0.4 and then continue at 18 °C. 

SECTION: Purification procedure
3. Once the cells reach an OD600 of 0.8, induce the protein expression with 100 micromolar (µM) isopropyl β-D-1-thiogalactopyranoside (IPTG) for 960 min at 18 °C.

SECTION: Purification procedure
7. Filter the supernatant through an 0.45 µm filter and load onto a pre-equilibrated 5 ml His-Trap HP column (Cytiva). 

SECTION: Purification procedure
6. Clear the lysates by centrifugation at 18000 rpm, 45 min, 4 °C  in a SORVAL RC6+ centrifuge with an F21S-8x50Y rotor (Thermo Scientific). 

SECTION: Purification procedure
5. Sonicate cell lysates twice for 30 s. 

SECTION: Purification procedure
4. Collect the cells by centrifugation and resuspend in lysis buffer, complete EDTA-free protease inhibitors (Roche), CIP protease inhibitor (Sigma), and DNase (Sigma)). 

Lysis buffer:
 
 
SECTION: Purification procedure
13. Pool the fractions containing purified mCherry-WIPI2d or mCherry-WIPI3. 

SECTION: Purification procedure
12. Analyse the fractions by SDS-PAGE and Coomassie staining. 

SECTION: Purification procedure
11. Elute the proteins with SEC buffer. 
 
SEC buffer:
 
 
SECTION: Purification procedure
10. Pool the fractions containing the 6xHis-TEV-mCherry-WIPI2d/WIPI3, concentrate using a 30 kDa cut-off Amicon filter (Merck Millipore) and loade onto a pre-equilibrated Superdex 200 Increase 10/300 GL column (Cytiva). 

SECTION: Purification procedure
9. Elute the proteins with a stepwise imidazole gradient (30, 75, 100, 150, 225, 300 mM). 

SECTION: Purification procedure
8. After bind His tagged proteins to the column, wash the column with three column volumes of wash buffer. 

Wash buffer:
 
 
SECTION: Purification procedure
15. Store the proteins at -80 °C.

SECTION: Purification procedure
14. After concentrating the purified protein, aliquot the protein and snap-frozen in liquid nitrogen.
TITLE: Intestinal Organoid Dissociation and Nuclei Isolation for Single Cell ATAC-Seq
AUTHORS: Heather Eckart, Ran Zhou, Nadia Khan

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This protocol provides a procedure for human intestinal organoid dissociation into a single cell suspension and nuclei isolation prior to Single Cell ATAC-Sequencing. </div></div>

[STEPS]
?. [Organoid Dissociation]
Incubate organoid in TrypLE for up to 20 minutes at 37°C .
?. [Nuclei Isolation]
For freshly dissociated cells, perform 1-2 washes with PBS + 0.04% BSA (20ul BSA/1mL 1X PBS).
Consult 10X Genomics protocol for using frozen cells.
?. [Nuclei Isolation]
Determine the cell count after washing  using a hemocytometer.
?. [Nuclei Isolation]
Add cell suspension of 100,000-1,000,000 cells to a 2-ml tube. For our experiment we started with 200,000 cells per sample.
?. [Organoid Dissociation]
Every 5 minutes, pipette the cell suspension up and down 5-10 x and check the digestion progress with a hemocytometer until over 90% of the cells are singlet cells.
?. [Nuclei Isolation]
Centrifuge at 300 rcf for 5 min at 4°C.
Centrifuge: 300 34, 5 min, 4 10
?. [Nuclei Isolation]
Remove ALL the supernatant without disrupting the cell pellet.
?. [Nuclei Isolation]
Add 100 µl chilled Lysis Buffer. Pipette to mix 10x.
[Lysis Buffer]
?. [Nuclei Isolation]
Incubate for 4 min on ice.
[4 min]
Time may vary depending on cell type; 4 minutes is specific for organoid samples.
?. [Nuclei Isolation]
Add 1 ml chilled Wash Buffer to the lysed cells. Pipette to mix 5x.
[Wash Buffer]
?. [Nuclei Isolation]
Centrifuge at 500 rcf for 5 min at 4°C.
Centrifuge: 500 34, 5 min, 4 10
?. [Nuclei Isolation]
Remove the supernatant without disrupting the nuclei pellet.
?. [Nuclei Isolation]
Based on your targeted nuclei recovery, cell concentration in step 4 and assuming ~50% nuclei loss during cell lysis, resuspend in chilled Diluted Nuclei Buffer (1x).  Maintain on ice.(See Nuclei Stock Concentration Table and Example Calculation below)For our experiment, we targeted 5,000 nuclei.
?. [Nuclei Isolation]
Check nuclei integrity by staining with WGA and DAPI. Also determine the nuclei concentration using a hemocytometer
?. [Nuclei Isolation]
OPTIONAL: If cell debris and large clumps are observed, pass through a cell strainer. For low volume, use a 40 µm Flowmi Cell Strainer to minimize volume loss
?. [Nuclei Isolation]
Proceed immediately to Chromium Next GEM Single Cell ATAC Sequencing protocol (found in the Chromium Single Cell ATAC Solution User Guide)
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p style="text-align: justify; text-indent: 1.0cm; line-height: 200%;"><span style="font-family: 'Times New Roman','serif'; color: black;">The novel c.804delG mutation, occurring in the seventh exon of the <em>ILDR1 </em>gene, abolishes a FauI restriction site. The FauI restriction pattern of the exon7 fragment (1003 bp) was used to screen 50 deaf individuals and 120 unrelated healthy UAE individuals. Digestion of PCR products was performed according to manufacturer’s instructions (New England Biolabs, USA), followed by separation on 2% agarose gels.</span></p>

[STEPS]
TITLE: Connecting     HTC Vive in SV location
AUTHORS: Ryan  Straight

[DESCRIPTION]
Connecting the HTC Vive for VR play while mirrored on television.

[BEFORE_START]
Double check that the Titan workstation is not reserved by anyone else.

[STEPS]
SECTION: Laptop setup
1. Save all work on the laptop.
SECTION: Laptop setup
2. Restart the laptop.
SECTION: Clean headset
3. While the laptop is restarting, get the corded HTC Vive from the display.
5. Log into Windows.
SECTION: Television
6. Turn on television.
SECTION: Television
8. Right-click on the speaker icon and choose "Sound settings."
SECTION: Television
9. Ensure "SAMSUNG TV" is selected.
SECTION: Connect headset
10. Attach HTC Vive to Titan laptop.
SECTION: Connect headset
11. Take the headset out of the Cleanbox and place it on the table next to the Titan laptop.
SECTION: Connect headset
12. Using the Vive 3-in-1 cable (tips ringed with orange), connect to the HTC Vive Link Box attached to the laptop.
SECTION: Connect headset
13. Grab the controllers off the charger and place next to the headset, ensuring they're in line-of-sight of the base stations.
SECTION: Steam
14. Start Steam on the Titan laptop.
SECTION: Steam
15. Close any pop-ups that may have opened (Steam will typically show an ad after starting).
SECTION: Steam
16. Ensure the mavrxlab Steam account is signed in.
SECTION: Steam
17. Make sure the SteamVR icons are lit up: headset, two controllers, and two sensors.

  

SECTION: Steam
18. Select Library in Steam, choose the program you wish to run, and hit the Play button.
SECTION: Television
7. Set volume.
SECTION: Clean headset
4. Use the Cleanbox protocol on the headset and controllers.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
This tutorial shows you how to create a pipeline on the University of Arizona HPC.  Specifically, we will be creating a pipeline to run blast, using scripts you are writing or have already written from ABE 487 weeks <a href="https://github.com/kyclark/abe487/tree/master/lab/week10" target="_blank">10 </a>and <a href="https://github.com/kyclark/abe487/tree/master/lab/week11" target="_blank">11</a>.  The pipeline will do the following:<br />1.  split each of the files from the night/day transcriptome into files with 10,000 sequences each<br />2.  run blastp against <span class="s1"> uniprot_sprot.fasta<br /></span>3.  parse the blast results<br /><br />

[GUIDELINES]
Before starting this tutorial, you need to complete the scripts from the computational homework for week <a href="https://github.com/kyclark/abe487/blob/master/lab/week10/problems.md" target="_blank">10</a> and <a href="https://github.com/kyclark/abe487/blob/master/lab/week10/problems.md" target="_blank">11</a>.<br /><br />

[STEPS]
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TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>Method for isolating mesophyll protoplasts from Arabidopsis as a system for monitoring RNA stability using transcriptional inhibitors (e.g. cordycepin) under stress treatments (e.g. high-light or hydrogen peroxide treatment).</p>
<p> </p>
<p>Adapted from Yoo S.-D., Cho Y.-H. &amp; Sheen J. (2007) Arabidopsis mesophyll protoplasts: a versatile cell system for transient gene expression analysis. Nature Protocols 2, 1565–1572.</p>

[BEFORE_START]
<p>Before starting experiment, it is considered ideal to keep intended <em>Arabidopsis</em> plants for protoplast extraction in the dark overnight. This is so that all the starch is used up, making the isolation procedure easier (less junk in extract).</p>
<p> </p>
<p>Prepare the following solutions:</p>
<p><strong>Enzyme Solution:</strong> Make solution of 20mM MES (pH 5.7), 0.4M Mannitol and 20mM KCl. Heat solution to 70°C for 3-5min. To this add cellulose R10 (1.5%) and macerozyme R10 (0.4%). Warm solution to 55°C for 10 min (inactivates DNAse and proteases and aids solubilising enzyme). Cool to room temperature and add 10mM Ca Cl<sub>2</sub>, 1-5mM ß-mercaptoethanol (optional) and 0.1% BSA. Filter the final enzyme solution through a 0.45um syringe filter into a petri dish (enzyme solution should be made fresh, however, can be stored overnight at 4°C).</p>
<p><strong>W5 Solution:</strong> 2mM MES (pH 5.7), 154mM NaCl, 125mM CaCl<sub>2</sub> and 5mM KCl.</p>
<p><strong>MMG Solution: </strong>4mM MES (pH5.7), 0.4M Mannitol and 15mM MgCl<sub>2</sub>.</p>

[GUIDELINES]
<p>Adapted from Yoo S.-D., Cho Y.-H. &amp; Sheen J. (2007) Arabidopsis mesophyll protoplasts: a versatile cell system for transient gene expression analysis. Nature Protocols 2, 1565–1572.</p>
<p> </p>
<p>Ideally, preps will be done on 3-4 week old <em>Arabidopsis </em>plants (before flowering). Choose well-expanded leaves (usually #5 &amp; #7).</p>
<p> </p>
<p>Protoplasting 80 leaves (using ~ 65 - 75 mL enzyme solution) from 3 week old plants yielded approx. 1.5 million protoplasts (diluted to 20,000/mL) from which 1.4 - 2.8 ug total RNA (of high quality) could be extracted using TRIzol.</p>
<p> </p>
<p>Higher RNA yields were obtained using fewer leaves and more enzyme solution (40-50 leaves of 3 week old plants gave ~4 ug total RNA).</p>
<p> </p>
<p> </p>

[STEPS]
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TITLE: No Title
AUTHORS: 

[DESCRIPTION]
This mixture is used in the <a href="https://www.protocols.io/view/CsCl-Step-Gradient-to-Purify-Phage-c4zyx5?step=4" target="_blank">CsCl Step Gradient to Purify Phage Protocol<br /></a>

[GUIDELINES]
<br /><strong>Grams of CsCl to be added to Buffer</strong><br /><br />
<table style="height: 223px;" width="472">
<tbody>
<tr>
<td>p</td>
<td>M</td>
<td>25 mls</td>
<td>50 mls</td>
<td>75 mls</td>
<td>100 mls</td>
<td>200 mls</td>
</tr>
<tr>
<td>1.3</td>
<td>2.4</td>
<td>10.10</td>
<td>20.20</td>
<td>30.31</td>
<td>40.41</td>
<td>80.82</td>
</tr>
<tr>
<td>1.4</td>
<td>3.2</td>
<td>13.47</td>
<td>26.94</td>
<td>44.16</td>
<td>53.86</td>
<td>107.76</td>
</tr>
<tr>
<td>1.45</td>
<td>3.6</td>
<td>&nbsp;</td>
<td>30.24</td>
<td>&nbsp;</td>
<td>60.48</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>1.5</td>
<td>4</td>
<td>16.87</td>
<td>33.74</td>
<td>50.61</td>
<td>67.48</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>1.55</td>
<td>4.4</td>
<td>18.48</td>
<td>36.96</td>
<td>&nbsp;</td>
<td>73.92</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>1.6</td>
<td>4.8</td>
<td>20.2</td>
<td>40.40</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>1.65</td>
<td>5.2</td>
<td>21.89</td>
<td>43.78</td>
<td>&nbsp;</td>
<td>87.55</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>1.7</td>
<td>5.6</td>
<td>23.57</td>
<td>47.15</td>
<td>70.7</td>
<td>94.29</td>
<td>&nbsp;</td>
</tr>
</tbody>
</table>

[STEPS]
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TITLE: C4 ZipTip Solid Phase Extraction 
AUTHORS: Lauren  Adams

[DESCRIPTION]
Solid phase extraction for clean-up and concentration of proteins prior to introduction into the mass spectrometer. 

[STEPS]
1. Activate Ziptip by pipetting 10 µL of C4 ZipTip Activation Buffer and discarding onto a Kimwipe for a total of 6 times.
2. Equilibrate the Ziptip by pipetting 10 µL of C4 ZipTip Equilibration/Wash Buffer and discarding onto a Kimwipe for a total of 6 times.
3. Remove C4 Ziptip from p20 pipette and place safely back into tip box to hold. Take a p200 pipette set at 200 µL and add a p200 pipette tip to the end. Take the p200 with pipette tip and then add the C4 ZipTip to the end. Carefully pipette the elution sample up and down for a total of 10 times per elution fraction. Avoid forcing air bubbles through the pipette tip as this will disrupt the resin and introduce oxidation to the target protein.
4. Remove C4 Ziptip from p200 pipette and place safely back into tip box to hold. Reattach to the p20 pipette and by pipet 10 µL of C4 ZipTip Equilibration/Wash Buffer and discard onto a Kimwipe for a total of 10 times.
5. Pipette into 5 µL of C4 ZipTip Elution Buffer that is inside a clean LoBind tube a total of 10 times. Dilute the final volume up to 25 µL for LC-MS or 80 µL for I2MS.
TITLE: Case Selection
AUTHORS: Chase Carver

[DESCRIPTION]
This protocol provides the basis for aged mouse acquisition and selection for brain tissue collection.

[STEPS]
SECTION: Case selection of aged mice
1. C57BL/6 Mice were acquired from the National Institute of Aging between 20-22 months old. The mice were housed in the vivarium for at least 2 weeks prior to tissue collection
Mice that demonstrated severe dermatitis were excluded from the study. Mice that exhibit overt splenomegaly at the time of necropsy are excluded from further analysis. Age at time of necropsy was 23-24 months old.
TITLE: Gait analysis using augmented reality markers
AUTHORS: Gergely Nagymáté, Rita M Kiss

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">The protocol provides guidelines, preparation and execution instructions to perform and process a gait analysis trial by a novel motion capture method using virtually calibrated anatomical points tracked by augmented reality (AR) markers.</div><div style = "text-align :; float : ;"><img style = "" src = "https://s3.amazonaws.com/pr-journal/ywxjg7w.gif" /></div></div>

[STEPS]
?. Attach the AR markers on the corresponding body segments (for gait analysis: pelvis, thighs and legs)
?. Start recording and perform anatomical calibration on the subject using the pointer after palpating the calibrated anatomical points.
?. Record the gait trial.
?. Upload the video takes to a computer.
?. The calibration and gait trial video files are processed offline by the image processing software where the frames of pointing to anatomical landmarks are selected manually.
?. After this manual post processing a file is available with the calculated marker trajectories during the trial in a standard .trc file format.
?. A third party open-source biomechanical analyzing software (OpenSim) can be used to calculate gait parameters.
TITLE: Streptavidin immovilization stress test
AUTHORS: Jorge Fernández

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">The following protocol was performed to test the affinity of the protein streptavidin for the nitrocellulose when applied a continuous flow of solvent.</div></div>

[STEPS]
?. Cut the FF80HP nitrocellulose membranes in  2 rectangles of 0.8cm x 4 cm dimensions. Repeat this procedure with the FF170HP nitrocellulose to obtain a total of 4 nitrocellulose pieces.
?. Stamp  the microfluidic design, following the protocol microfluidic channels wax priming on:One piece of the FF170Hp nitrocellulose membraneOne piece of the FF80HP nitrocellulose membrane.
?. Glue the absorbant pad with tape at the top edge top of the 4 pieces of nitrocellulose. the dimensions of the absorbant pad are 0.8 cm x 1.2 cm.
?. Deposit 0,33 ul of streptavidin solution with a micropipette. Repeat this procedure 3 times, letting the drop dry in between each time.The procedure must be done in the 4 pieces of nitrocellulose.
?. Let the pieces dry overnight at 4ºC in a desiccator.
?. Perform a thin-layer chromatography with PBS-Tween as the solvent: pour 15 ml of the solvent and position the pieces with the absorbant touching the bottom of the container. Let the solvent flow trough the nitrocellulose for 15 min.The assay can be done individually or with the 4 pieces of nitrocellulose at one time.
?. Let it dry at room temperature for 40 min
?. Reveal with  Ponceau 0.1% within acetic acid 5% by immersing the nitrocellulose piece into the solution for a few seconds.
?. Destain in dH2O in agitation for 2 min.