Name: antifouling self-assembled monolayers on microelectrodes for patterning biomolecules
Uploaded: 2009-08-25T16:00:00
Description: Watch this Scientific Journal Video about Antifouling Self-assembled Monolayers on Microelectrodes for Patterning Biomolecules at JoVE.com

<p class="jove_content">We acknowledge Melissa Grunlan for helpful discussions on SAM formation. This research was supported in part by the Robert A. Welch Foundation (A-1585).</p>

<p class="jove_title">Preparation of reagents</p>
<p class="jove_content">Prepare PIPES buffer (80 mM PIPES, 1mM MgCl<sub>2</sub>, 1mM EGTA, adjusted to pH 6.8 with KOH). Aliquot and store glucose oxidase, catalase and glucose at -70&deg;C and taxol at -20&deg;C according to existing protocols.<sup>1,2</sup> Aliquot and store tubulin at -70&deg;C according to the instructions included by the supplier.</p>
<p class="jove_title">Pattern Au microelectrodes using lithography</p>
<ol>
<li>Cut silicon wafers into 1 cm &acute; 1.5 cm substrates using ...

<ol>
	<li>Rai-Choudhury, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> <em style='font-style: italic'>Handbook of Microlithography, Micromachining, and Microfabrication: Microlithography</em>. , (1997).</li></ol>

<p class="jove_content">The migration of MTs is easily visualized using fluorescence microscopy because of their brightness and the low background fluorescence. The method is generally applicable to biomolecule patterning, as it only requires that the biomolecules be induced to carry a net charge by suitable adjustment of the buffer pH. Electroosmosis is avoided in this setup because there are no charged surfaces such as the silica walls used in capillary electrophoresis.</p>
<p class="jove_content">Several modifications of this method are pos...

<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">	
		
		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>2-[methoxypoly-(ethyleneoxy)propyl]-trimethoxysilane (PEG-silane)</td>
<td>&nbsp;</td>
<td>Gelest, Inc.</td>
<td>SIM6492.7</td>
<td>6-9 PEG units;
molecular weight 450-600 g mol<sup>-1</sup></td>
</tr>
<tr>
<td>2-mercaptoethanol</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>M7522</td>
<td>&nbsp;</td>
<tr>
<td>Acetone</td>
<td>&nbsp;</td>
<td>EMD Chemicals</td>
<td>AX0120-8</td>
<td>ACS grade</td>
</tr>
<tr>
<td>Catalase</td>
<td>&nbsp;</td>
<td>Calbiochem</td>
<td>219001-5MU</td>
<td>&nbsp;</td>
<tr>
<td>Chlorobenzene</td>
<td>&nbsp;</td>
<td>EMD Chemicals</td>
<td>MK441908</td>
<td>ACS grade</td>
</tr>
<tr>
<td>Ethylene glycol-bis(2-aminoethylether)<em>-N,N,N&#x2019;,N&#x2019;-</em>tetraacetic acid (EGTA)</td>
<td>&nbsp;</td>
<td>Calbiochem</td>
<td>324626</td>
<td>&nbsp;</td>
<tr>
<td>Glucose</td>
<td>&nbsp;</td>
<td>EMD Chemicals</td>
<td>DX0145-1</td>
<td>&nbsp;</td>
<tr>
<td>Glucose oxidase</td>
<td>&nbsp;</td>
<td>MP Biomedicals</td>
<td>100330</td>
<td>&nbsp;</td>
<tr>
<td>Guanosine-5&#x2019;-[(&alpha;,&Beta;)-methyleno]triphosphate, Sodium salt (GMPCPP)</td>
<td>&nbsp;</td>
<td>Jena Biosciences, GmbH</td>
<td>NU-405s</td>
<td>&nbsp;</td>
<tr>
<td>Guanosine-5&#x2019;-triphosphate</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>G8752</td>
<td>&nbsp;</td>
<tr>
<td>Isopropanol</td>
<td>&nbsp;</td>
<td>EMD Chemicals</td>
<td>PX1835-5</td>
<td>ACS grade</td>
</tr>
<tr>
<td>Methyl isobutyl ketone (MIBK)</td>
<td>&nbsp;</td>
<td>Fisher Scientific</td>
<td>AC12739-0010</td>
<td>&nbsp;</td>
<tr>
<td>Paclitaxel (taxol)</td>
<td>&nbsp;</td>
<td>Calbiochem</td>
<td>580555</td>
<td>&nbsp;</td>
<tr>
<td>Piperazine-N,N''-bis(2-ethanesulfonic acid) (PIPES)</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>P1851</td>
<td>&nbsp;</td>
<tr>
<td>Polymethylmethacrylate (PMMA)</td>
<td>&nbsp;</td>
<td>Brewer Science, Inc.</td>
<td>950K</td>
<td>molecular weight 950K</td>
</tr>
<tr>
<td>Rhodamine tubulin</td>
<td>&nbsp;</td>
<td>Cytoskeleton, Inc.</td>
<td>T331M</td>
<td>from bovine brain 99% pure, lyophilized</td>
</tr>
<tr>
<td>Toluene</td>
<td>&nbsp;</td>
<td>EMD Chemicals</td>
<td>TX0735-5</td>
<td>ACS grade</td>
</tr>
<tr>
<td>Tubulin</td>
<td>&nbsp;</td>
<td>Cytoskeleton, Inc.</td>
<td>T238</td>
<td>from bovine brain 99% pure, lyophilized</td>
</tr>		</tbody>
		</table>
		</div>
			<div>
			<p class="jove_content">No. 1 borosilicate glass coverslips (22 X 50 mm) were purchased from VWR. Polished electronic grade p-type &lt;111&gt; silicon wafers were purchased from Addison Engineering, Inc. High-purity silver paint was obtained from Structure Probe, Inc. (Catalog # 5001 AB). Insulated fine gauge copper magnet wire (~ 0.08 mm (0.003&#148;) diameter) can be obtained from electronic supply companies.</p>		</div>
		

antifouling self-assembled monolayers on microelectrodes for patterning biomolecules

We present a procedure for forming a poly(ethylene glycol) (PEG) trimethoxysilane self-assembled monolayer (SAM) on a silicon substrate with gold microelectrodes. The PEG-SAM is formed in a single assembly step and prevents biofouling on silicon and gold surfaces. The SAM is used to coat microelectrodes patterned with standard, positive-tone lithography. Using the microtubule as an example, we apply a DC voltage to induce electrophoretic migration to the SAM-coated electrode in a reversible manner. A flow chamber is used for imaging the electrophoretic migration and microtubule patterning <em>in situ</em> using epifluorescence microscopy. This method is generally applicable to biomolecule patterning, as it employs electrophoresis to immobilize target molecules and thus does not require specific molecular interactions. Further, it avoids problems encountered when attempting to pattern the SAM molecules directly using lithographic techniques. The compatibility with electron beam lithography allows this method to be used to pattern biomolecules at the nanoscale.

Watch this Scientific Journal Video about Antifouling Self-assembled Monolayers on Microelectrodes for Patterning Biomolecules at JoVE.com

Antifouling Self-assembled Monolayers on Microelectrodes for Patterning Biomolecules

<p class="jove_content">We present a procedure for forming a poly(ethylene glycol) self-assembled monolayer (PEG-SAM) on a silicon substrate with gold microelectrodes. The PEG-SAM is formed in a single step and prevents biofouling on silicon and gold surfaces. Electrophoresis is then used for patterning biomolecules down to the nanoscale.</p>

We present a procedure for forming a poly(ethylene glycol) (PEG) trimethoxysilane self-assembled monolayer (SAM) on a silicon substrate with gold ...

Research

JoVE Journal

Biology

Patterning Cells on Optically Transparent Indium Tin Oxide Electrodes

The ability to exercise precise spatial and temporal control over cell-surface interactions is an important prerequisite to the assembly of multi-cellular ...

Automated System for Single Molecule Fluorescence Measurements of Surface-immobilized Biomolecules

Fluorescence Resonance Energy Transfer (FRET) microscopy has been widely used to study the structure and dynamics of molecules of biological interest, ...

Single Cell Transcriptional Profiling of Adult Mouse Cardiomyocytes

While numerous studies have examined gene expression changes from homogenates of heart tissue, this prevents studying the inherent stochastic variation ...

Protein WISDOM: A Workbench for <em>In silico</em> <em>De novo</em> Design of BioMolecules

Protein WISDOM: A Workbench for In silico De novo Design of BioMolecules

The aim of de novo protein design is to find the amino acid sequences that will fold into a desired 3-dimensional structure with improvements in specific ...

Internalization and Observation of Fluorescent Biomolecules in Living Microorganisms via Electroporation

The ability to study biomolecules in vivo is crucial for understanding their function in a biological context. One powerful approach involves fusing ...

Measurement and Analysis of Extracellular Acid Production to Determine Glycolytic Rate

Extracellular measurement of oxygen consumption and acid production is a simple and powerful way to monitor rates of respiration and glycolysis1. Both ...

Isolation of Specific Genomic Regions and Identification of Associated Molecules by enChIP

The identification of molecules associated with specific genomic regions of interest is required to understand the mechanisms of regulation of the ...

Quantification of Site-specific Protein Lysine Acetylation and Succinylation Stoichiometry Using Data-independent Acquisition Mass Spectrometry

Post-translational modification (PTM) of protein lysine residues by N&#400;-acylation induces structural changes that can dynamically regulate protein ...

SA-&#946;-Galactosidase-Based Screening Assay for the Identification of Senotherapeutic Drugs

Cell senescence is one of the hallmarks of aging known to negatively influence a healthy lifespan. Drugs able to kill senescent cells specifically in cell ...

Simultaneous Affinity Enrichment of Two Post-Translational Modifications for Quantification and Site Localization

Studying multiple post-translational modifications (PTMs) of proteins is a crucial step to understand PTM crosstalk and gain more holistic insights into ...