This work was supported by a grant from the NIH. The assays shown in Figure 1 was performed by Dr. Joseph Fasolo. We thank Dr. Rui Chen for helpful comments.

1. Probe Preparation
<ol>
	<li>Culture and purify the V5-fusion kinase probes used to examine interactions with other proteins as follows:
	<ol>
		<li>Use freshly streaked yeast strain Y258 (MATa pep4-3, his4-580, ura3-53, leu2-3,112) containing V5-fusion protein (expressed from GATEWAY vector pYES-DEST52). Use the isolated protein as probe on the microarrays. Plate the yeast on synthetic complete-uracil (Sc-Ura)/2% Dextrose/Agar and grow at 30&#160;&#176;C&#160;for 3 days from frozen culture (-80 &#176;C glycerol stock).</li>
		<li>Inoculate starter cultures (5&#8211;20 ml) from a single colony and grow overnight in Sc-Ura/2% Dextrose on shaking platform (220-250 rpm) or wheel at 30&#160;&#176;C.</li>
		<li>The following morning, inoculate 400 ml&#160;of the Sc-Ura/2% raffinose culture with sufficient starter culture to a final OD600 of 0.1.</li>
		<li>Grow the inoculums to OD600 of 0.6 followed by galactose induction of the V5-kinase fusion construct expression by adding a solution of 3x&#160;Yeast Extract/Peptone (YEP) supplemented with 6% Galactose, by adding enough to dilute the induction media by a factor of 3 so that final concentration of galactose is 2%.</li>
		<li>Induce cells at 30 &#176;C for 6 hr&#160;on a shaking platform. Use a 2 L&#160;Erlenmeyer flask to ensure appropriate aeration.</li>
		<li>Harvest cells using a JA-10 (or comparable) rotor by spinning 400 ml&#160;of cell suspension at 1,000 x g for 5 min at 4 &#176;C.</li>
		<li>Wash the cells once with 50 ml&#160;of ice cold PBS buffer&#160;and transfer to a 50 ml&#160;conical tube. Wash the pellet again in ice cold PBS buffer (without detergents or other additives) used for lysis and transfer to 2 ml&#160;snap-cap tubes for lysis.</li>
		<li>Spin the cells at 20,000 x g for 1 min at 4 &#176;C to a pellet and pipette away the buffer.</li>
		<li>Place tubes on ice and proceed with lysis step.</li>
		<li>Lyse the cells (250-350 &#181;l pellet) with 0.5 mm zirconia beads in a 1:1:1 volume of cell pellet, beads, and Phosphate-buffered saline ( PBS) lysis buffer and vortex the mixture using a agitation platform 3 times at 2 min intervals at 4 &#176;C.</li>
		<li>Centrifuge the lysate at 20,000 x g in a tabletop microfuge for 10 min at 4 &#176;C.</li>
		<li>Pipette supernatant into polycarbonate high speed centrifuge tube and clarify by ulracentrufugation for 30 min at 150,000 x g at 4 &#176;C.</li>
		<li>Transfer the clarified lysate to a tube containing prewashed Ni2+ affinity resin (~100 &#181;l) and incubate on a nutator for 2 hr&#160;at 4 &#176;C to capture Histidine(His)6X tagged V5-fusion protein.</li>
		<li>Wash the resin three times for 10 min&#160;at 4 &#176;C with Wash Buffer. Pellet the resin using a table top centrifuge for 5 min at 1,000 x g at 4 &#176;C and aspirate the supernatant. Add fresh wash buffer for each wash and return the tube the nutator for agitation during the wash.</li>
		<li>After the wash steps are complete, apply the washed resin to a fresh G-25 column for elution.</li>
		<li>Apply 25 &#181;l of elution buffer in a step-wise manner beginning with 100 mM Imidazole to 500 mM imidazole in 50 mM increments for a total of 9 fractions.</li>
		<li>Assay the collected fractions using sodium dodecyl sulfate polyacrylamide gel (SDS-PAGE) and coomassie staining to identify the most highly enriched fraction(s) for use in the assay and add glycerol to 30% and store at -80&#160;&#176;C&#160;until used in the assay.</li>
	</ol>
	</li>
</ol>
2. Probing the Arrays
NOTE: Please see step 2.6 of this section to prepare the antibody solution before beginning the assay.
<ol>
	<li>To detect interactions, dilute the V5-fluorophore conjugated antibody (i.e. AlexaFluor 647 ) to 260 ng/ml&#160;in probe buffer and mix thoroughly by shaking. 
	NOTE: Prepare this antibody solution 30 min&#160;prior to use and place the tube on a nutator or wheel to ensure complete mixing and a homogenous suspension of antibody.</li>
	<li>Dilute the V5-fusion protein probe over a concentration range of 5&#8211;500 &#956;g/ml. 
	NOTE: Optimized for each protein&#8211;protein interaction assay, using probe buffer. Optimization involves adding more probes to probe buffer. Typically 10 &#181;g/ml is used as starting point and adjusted accordingly based on the signal strength.</li>
	<li>Remove the protein microarrays from the freezer (&#8722;20 &#176;C) and bring to 4 &#176;C in the refrigerator just prior to use.</li>
	<li>Add blocking buffer directly to the slide holder containing the protein micorarrays and cover the top with parafilm to prevent leakage. Block the arrays in blocking buffer for 1 hr&#160;by shaking at 50 rpm on a stage at 4 &#176;C.</li>
	<li>After blocking, transfer the arrays to a humidified chamber chilled to 4&#160;&#176;C, and add 90 &#956;l&#160;of diluted probe directly to the array surface. Overlay the arrays with a raised lifter slip and incubate static (no shaking) in the humidified chamber at 4&#160;&#176;C&#160;for 1.5 hr.</li>
	<li>Wash the arrays 3 times for 1 min each in probe buffer in three 50 ml&#160;conical tubes. Add the slide to conical tubes containing enough pre-chilled probe buffer to completely envelope the slide. Allow the lifter slip to gently slide off of the protein microarray (do not force it off as this could result in damage to the array surface).</li>
	<li>Apply the antibody solution directly to array immediately after completing the wash (step 2.5) and overlay with a raised lifter slip as before. Incubate the arrays for 30 min at 4 &#176;C in the humidified chamber.</li>
	<li>Perform the same wash step as before (3 times 1 min in probe buffer), and spin in a 50 ml&#160;conical tube at 800 x g in a tabletop centrifuge for 5 min at room temperature. Air-dry the arrays in a slide holder in the dark for 30 min prior to scanning the array at 647 nm.</li>
</ol>

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C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+organization+of+the+yeast+proteome+by+systematic+analysis+of+protein+complexes.">Functional organization of the yeast proteome by systematic analysis of protein complexes.</a> Nature. 415, 141-147 (2002).</li><li>Jeong, J. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+identification+of+monospecific+monoclonal+antibodies+using+a+human+proteome+microarray.">Rapid identification of monospecific monoclonal antibodies using a human proteome microarray.</a> Mol Cell Proteomics. 11 (6), (2012).</li><li>Krogan, N. 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C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+binding+of+calmodulin-related+proteins+to+their+targets+revealed+through+high-density+Arabidopsis+protein+microarrays.">Differential binding of calmodulin-related proteins to their targets revealed through high-density Arabidopsis protein microarrays.</a> Proceedings of the National Academy of Sciences of the United States of America. 104, 4730-4735 (2007).</li><li>Zhu, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Global+analysis+of+protein+activities+using+proteome+chips.">Global analysis of protein activities using proteome chips.</a> Science. 293, 2101-2105 (2001).</li><li>Ptacek, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Global+analysis+of+protein+phosphorylation+in+yeast.">Global analysis of protein phosphorylation in yeast.</a> Nature. 438, 679-684 (2005).</li><li>Haab, B. 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The authors declare that they have no competing financial interests.

The protocol presented was originally performed using 85 unique yeast protein kinase-V5 fusion proteins to compare binding activity across distinct and related families of yeast protein kinases resulting in the identification of new kinase interaction networks in vivo1. As an emerging proteomic profiling technology, the development of High Throughput (HTP) screening of proteomes using protein microarrays relied on peptide libraries, and eukaryotic and prokaryotic model organisms8,17; later,...

The need to perform global analyses of protein biochemistry and binding activity in vivo has resulted in the development of new methods for profiling protein-protein interactions (PPIs) and the post-translational modifications of whole proteomes1,3-8. Protein microarrays are manufactured as functional protein microarrays using full-length functional proteins4-6,8,9, or analytical protein microarrays containing antibodies10,11. They are engineered to contain a high-density of proteins arrayed onto microscope slides with a variety of surface chemistries to facilitate a variety of experimental conditions required for conducting wide-ranging biochemical analyses12. Nitrocellulose and aldehyde surface chemistries for chemical attachment through lysine or affinity attachment methods such as nickel- chelated slides for attaching His-tagged proteins and glutathione for affinity attachment among others13.
The use of functional protein microarrays to detect protein-protein interactions requires access to a high-quality functional protein library14. S. cerevisiae is amenable to producing such a library through the pairing of high-copy affinity tagged protein constructs with high-throughput chromatographic purification techniques. The vast majority of the yeast genome has been sequenced and nearly the entire proteome can be expressed from a high-copy plasmid for purification and biochemical analyses12. Once the proteins are obtained and arrayed in 384-well format, they are printed onto a microscope slide allowing for rapid parallel multi-parametric biochemical analysis and bioinformatic interrogation8,14-16. Protein microarrays have been used for enzymatic assays and interactions with proteins, lipids, small molecules, and nucleic acids among many other applications. The accessibility of proteins on the surface of proteome arrays make them amenable to different types of analytical detection including, immune-affinity, Surface Plasmon Resonance, fluorescence and many other techniques. Moreover, it allows for fine control of the experimental condition where it might be hard to do in vivo.
The aim of this protocol is to demonstrate the appropriate use of functional protein microarrays to detect protein-protein interactions. This application enables the high-throughput parallel biochemical analysis of protein binding activities using a highly purified analyte (protein) of interest. A C-terminal(carboxy-terminal) tagged V5-fusion bait protein of interest is produced from a high-copy plasmid in a yeast strain optimized for protein purification. C-terminal tagging ensures that the full-length protein has been translated. The protein used in this study is Tda1-V5 fusion protein kinase, which is purified using nickel affinity resin via a His6X tag. The Tda1-V5 fusion construct is purified through serial elution using an imidazole gradient to elute the most highly enriched fraction for use in the assay.

<table border="0" cellpadding="0" cellspacing="0" width="855">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Name of Material/ Equipment</td>
			<td style="width:180px;">Company</td>
			<td style="width:119px;">Catalog Number</td>
			<td style="width:291px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Petri Dishes</td>
			<td style="width:180px;">VWR</td>
			<td>NC-10747</td>
			<td>or comparable</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Bacto yeast extract</td>
			<td style="width:180px;">Difco</td>
			<td>0217-17</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Bacto peptone</td>
			<td style="width:180px;">Difco</td>
			<td>0118-17</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Bacto agar</td>
			<td style="width:180px;">Difco</td>
			<td>0140-01</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Dextrose</td>
			<td style="width:180px;">Sigma</td>
			<td>D9434</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Raffinose</td>
			<td style="width:180px;">Sigma</td>
			<td>R0514</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Galactose</td>
			<td style="width:180px;">Sigma</td>
			<td>G0750</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Sc-Ura drop out media</td>
			<td style="width:180px;">MP Bio</td>
			<td>114410622</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Small Culture tubes</td>
			<td style="width:180px;">VWR/Fisher</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Large Erlenmeyer Flask</td>
			<td style="width:180px;">VWR/Fisher</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Centrifuge JA-10 rotor</td>
			<td style="width:180px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Centrifuge tubes (500 mL)</td>
			<td style="width:180px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Falcon tube (50 mL)</td>
			<td style="width:180px;">VWR/Fisher</td>
			<td>21008-951</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">PBS Tablets</td>
			<td style="width:180px;">Sigma</td>
			<td>P4417</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">FastPrep Tubes(2ml screw cap tube)</td>
			<td style="width:180px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">FastPrep Machine</td>
			<td style="width:180px;">MP Biomedicals</td>
			<td align="right" style="width:119px;">116004500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Zircon Beads</td>
			<td style="width:180px;">BioSpec</td>
			<td align="right" style="width:119px;">11079105</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Triton X100</td>
			<td style="width:180px;">Sigma</td>
			<td>P4417</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">DTT</td>
			<td style="width:180px;">Sigma</td>
			<td>D9779</td>
			<td></td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:267px;">MgCl2</td>
			<td style="width:180px;">Sigma</td>
			<td>M8266&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">NaCl</td>
			<td style="width:180px;">Sigma</td>
			<td>S3014&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Imidazole (pH = 7.4)</td>
			<td style="width:180px;">Sigma</td>
			<td>I5513&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">PMSF</td>
			<td style="width:180px;">Sigma</td>
			<td>93482</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Complete Inhibitor Cocktail (EDTA Free)</td>
			<td style="width:180px;">Roche</td>
			<td style="width:119px;">11873580001</td>
			<td style="width:291px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Phosphatase inhibitor cocktail 1</td>
			<td style="width:180px;">sigma</td>
			<td>P2850</td>
			<td style="width:291px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">BSA</td>
			<td style="width:180px;">Sigma</td>
			<td>A2153&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Tween-20</td>
			<td style="width:180px;">Sigma</td>
			<td>P1379&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">ATP</td>
			<td style="width:180px;">Sigma</td>
			<td>A1852&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Shaking Incubator (30 &#176;C)</td>
			<td style="width:180px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Nickel Affinity Resin</td>
			<td style="width:180px;">Life Technologies</td>
			<td>R901-01</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">G25 column</td>
			<td style="width:180px;">GE life sciences</td>
			<td>27-5325-01&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Nutator</td>
			<td style="width:180px;">VWR/Fisher</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">SDS-PAGE Gel (NuPage)</td>
			<td style="width:180px;">Life Technologies</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Coomassie Blue Stain</td>
			<td style="width:180px;">Life Technologies</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Monoclonal V5 Antibody</td>
			<td style="width:180px;">Life Technologies</td>
			<td>R960-25</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:267px;">Alexa647 Tagged monoclonal V5 Antibody</td>
			<td style="width:180px;">Life Technologies</td>
			<td>451098</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Protein Microarrays Kit</td>
			<td style="width:180px;">Life Technologies</td>
			<td>PAH0525013</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:267px;">Genepix Scanner</td>
			<td style="width:180px;">Molecular Devices</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="62">
			<td height="62" style="height:63px;width:267px;">Lifter Slips</td>
			<td style="width:180px;">Thomas Scientific</td>
			<td style="width:119px;"></td>
			<td style="width:291px;">http://www.thomassci.com/Supplies/Microscope-Cover-Glass/_/LIFTER-SLIPS/</td>
		</tr>
		<tr height="124">
			<td height="124" style="height:124px;width:267px;">Lysis Buffer: 0.1% Triton X-100, 0.5 mM DTT, 
			2 mM MgCl2, 500 mM NaCl, 50 mM imidazole (pH 7.4), Complete protease inhibitor cocktail &#8211; EDTA-free, Phosphatase inhibitor Cocktail 1, 1 mM PMSF&#160; (add just prior to use)</td>
			<td style="width:180px;"></td>
			<td style="width:119px;"></td>
			<td style="width:291px;">Add the reagents to 1 X PBS (0.01 M phosphate buffer, 0.0027 M potassium chloride, 0.137 M sodium chloride; pH 7.4) to obtain the desired volume of lysis solution</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:267px;">Blocking Buffer: 1% bovine serum albumin (BSA), 0.1% Tween-20</td>
			<td style="width:180px;"></td>
			<td style="width:119px;"></td>
			<td style="width:291px;">Add the reagents to 1 X PBS to obtain the desired volume of blocking buffer</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:267px;">Probe Buffer: 2 mM MgCl2, 0.5 mM DTT, 0.05% Triton X-100, 50 mM NaCl,500 &#956;M ATP (for kinases), 1% BSA</td>
			<td style="width:180px;"></td>
			<td style="width:119px;"></td>
			<td style="width:291px;">Add the reagent to 1 X PBS to obtain desired volume of probe buffer</td>
		</tr>
		<tr height="67">
			<td height="67" style="height:67px;width:267px;">Wash Buffer: 0.1% Triton X-100, 500 mM NaCl, 0.5 mM DTT, 50 mM imidazole (pH 7.4),2 mM MgCl2</td>
			<td style="width:180px;"></td>
			<td style="width:119px;"></td>
			<td style="width:291px;">Add the reagent to 1 X PBS to obtain desired volume of probe buffer</td>
		</tr>
		<tr height="168">
			<td height="168" style="height:168px;width:267px;">Elution Buffer: 0.1% Triton X-100, 0.5 mM DTT, 2 mM MgCl2, 500 mM NaCl, 50 &#8211; 500 mM imidazole (pH 7.4), Complete protease inhibitor cocktail &#8211; EDTA-free, Phosphatase inhibitor Cocktail 1, 1 mM PMSF (add just prior to use)</td>
			<td style="width:180px;"></td>
			<td style="width:119px;"></td>
			<td style="width:291px;">Before you begin, it is important to note the concentration gradient of imidazole (50 &#8211; 500 mM) and to create separate tubes for each concentration (it is advisable to create the interval using 50 mM increments). Add the reagent to 1 X PBS to obtain desired volume of probe buffer</td>
		</tr>
		<tr height="105">
			<td height="105" style="height:105px;width:267px;">3x YEP +6% galactose: Dissolve 30 g of yeast extract and&#160; 60 g of peptone in 700 ml of H2O, and autoclave. Add 300 ml of filter sterilized 20% galactose (wt/vol)</td>
			<td style="width:180px;"></td>
			<td style="width:119px;"></td>
			<td>&#160;galactose should not be autoclaved</td>
		</tr>
	</tbody>
</table>

probing high-density functional protein microarrays to detect protein-protein interactions

High-density functional protein microarrays containing ~4,200 recombinant yeast proteins are examined for kinase protein-protein interactions using an affinity purified yeast kinase fusion protein containing a V5-epitope tag for read-out. Purified kinase is obtained through culture of a yeast strain optimized for high copy protein production harboring a plasmid containing a Kinase-V5 fusion construct under a GAL inducible promoter. The yeast is grown in restrictive media with a neutral carbon source for 6 hr followed by induction with 2% galactose. Next, the culture is harvested and kinase is purified using standard affinity chromatographic techniques to obtain a highly purified protein kinase for use in the assay. The purified kinase is diluted with kinase buffer to an appropriate range for the assay and the protein microarrays are blocked prior to hybridization with the protein microarray. After the hybridization, the arrays are probed with monoclonal V5 antibody to identify proteins bound by the kinase-V5 protein. Finally, the arrays are scanned using a standard microarray scanner, and data is extracted for downstream informatics analysis1,2 to determine a high confidence set of protein interactions for downstream validation in vivo.

The protein-protein interaction activity was observed using a standard chip reader to evaluate the Tda1-V5 protein kinase fusion construct as a bait protein against a yeast functional protein microarray containing approximately 4,200 unique S. cerevisiae GST-fusion proteins. Further interrogation with the Genepix software revealed a multitude of binding events of varying intensities. The affinity was gauged from the graded intensity of the signal derived from a monoclonal V5-Fluorophore conjugated antibody bound...

Watch this Scientific Journal Video about Probing High-density Functional Protein Microarrays to Detect Protein-protein Interactions at JoVE.com

Probing High-density Functional Protein Microarrays to Detect Protein-protein Interactions

Using protein microarrays containing nearly the entire S. cerevisiae proteome is probed for rapid unbiased interrogation of thousands of protein-protein interactions in parallel. This method can be utilized for protein-small molecule, posttranslational modification, and other assays in high-throughput.

High-density functional protein microarrays containing ~4,200 recombinant yeast proteins are examined for kinase protein-protein interactions using an ...

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JoVE Journal

Biology

7.9K Views.  Stanford University. Using protein microarrays containing nearly the entire S. cerevisiae proteome is probed for rapid unbiased interrogation of thousands of protein-protein interactions in parallel. This method can be utilized for protein-small molecule, posttranslational modification, and other assays in high-throughput.